The accessibility of the thiol groups on G- and F-actin of rabbit muscle.
AuthorsLiu DF, Wang D, Stracher A
JournalBiochem J
PubMed ID2317199
The accessibility of the cysteine residues of actin from rabbit muscles to the thiol-targeted reagent 7-dimethylamino-4-methyl-(N-maleimidyl)coumarin (DACM) was investigated. Under conditions where the actin is in the unpolymerized form (G-actin), the most reactive thiol group was Cys-257, suggesting that it was located on the surface of the actin molecule. The ... More
Fluorescence stopped-flow study of N-(7-dimethylamino-4-methyl-3-coumarinyl)-maleimide-labeled troponin C.
AuthorsIio T, Kondo H
JournalJ Biochem (Tokyo)
PubMed ID7451407
Static fluorescence titration and fluorescence stopped-flow kinetic experiments o N-(7-dimethylamino-4-methyl-3-coumarinyl)-maleimide-labeled troponin C showed that the molecular kinetic mechanism of the local conformational change induced by the rapid Ca2+ binding and removal reactions with the high affinity Ca2+-binding sites (sites III and IV) can be explained by the following scheme: [Formula: ... More
Interaction of the fluorescence-labeled secretory component with human polymeric immunoglobulins.
AuthorsGoto Y, Aki K
JournalBiochemistry
PubMed ID6529580
'The secretory component (SC) isolated from human milk was labeled with 2 mol of the fluorescent thiol reagent N-[7-(dimethylamino)-4-methylcoumarinyl]maleimide (DACM) per mol of SC through the reactive disulfide bond of SC. The binding of the labeled SC to polymeric immunoglobulins was examined by gel filtration by measuring the fluorescence of ... More
The histochemical distribution of protein bound sulfhydryl groups in human epidermis by the new staining method.
AuthorsOgawa H, Taneda A, Kanaoka Y, Sekine T
JournalJ Histochem Cytochem
PubMed ID90070
'Recently, we synthesized a new fluorescent thiol reagent, N-(7-dimethylamino-4-methylcoumarinyl)-maleimide (DACM) which is nonfluorescent by itself but will react readily with -SH groups to form highly fluorescent addition products. By the use of this reagent, we studied the localization and concentration of -SH groups and S--S linkages in the human epidermis. ... More
Differential staining of skin-limited amyloid and colloid bodies with immunofluorescence after pretreatments.
AuthorsMukai H, Eto H, Nishiyama S, Hashimoto K
JournalJ Invest Dermatol
PubMed ID2450933
'We examined whether immunoglobulin (Ig) and complement (C) components of amyloid and colloid bodies were inherent parts of these substances or were present due to nonspecific absorption only. In direct immunofluorescence (IF) studies without pretreatment, skin-limited and systemic amyloid and colloid bodies in all cases showed positive staining for Ig ... More
Reversed-phase ion-pair high-performance liquid chromatography of mercaptoacetate and N-acetylcysteine after derivatization with N-(1-pyrene)maleimide and N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide.
AuthorsKågedal B, Källberg M
JournalJ Chromatogr
PubMed ID7096475
'We have developed a high-performance liquid chromatographic system capable of resolving mercaptoacetate and N-acetylcysteine as their N-(1-pyrene)maleimide (PM) and N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide (DACM) derivatives. Good resolution was obtained by ion pairing with tetramethylammonium hydroxide and chromatography on reversed phase. The detection limits for the thiols were about 50 fmol as their DACM ... More
Imaging of cAMP-dependent protein kinase activity in living neural cells using a novel fluorescent substrate.
AuthorsHigashi H, Sato K, Ohtake A, Omori A, Yoshida S, Kudo Y
JournalFEBS Lett
PubMed ID9305731
'In order to visualize the activity of the cAMP-dependent protein kinase (PKA) in living cells, we have constructed a new fluorescence PKA substrate by conjugating a fluorescence probe to a partial amino acid sequence of PKA regulatory domain II which contains a specific autophosphorylation site. The fluorescent peptide was cell-permeable ... More
A high-sensitivity fluorometric high-performance liquid chromatographic method for determination of glutathione and other thiols in cultured melanoma cells, microdialysis samples from melanoma tissue, and blood plasma.
AuthorsDizdar N, Kågedal B, Smeds S, Arstrand K
JournalMelanoma Res
PubMed ID1822768
'A high-performance liquid chromatographic method with fluorometric detection is described which is suitable for determination of glutathione in small samples. Reduced glutathione (GSH) and total glutathione obtained as GSH after reduction with glutathione reductase is derivatized with N-(7-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM) and subjected to chromatography. The detection limit for the GSH-DACM ... More
Sulfhydryl and disulfide stainings in amyloids of skin-limited and systemic amyloidoses.
AuthorsMukai H, Kanzaki T, Nishiyama S
JournalJ Invest Dermatol
PubMed ID6197491
'Disulfide (S-S) bonds and sulfhydryl (-SH) groups in skin-limited and systemic amyloidoses in frozen and paraffin-embedded sections were examined with a thiol reagent, N-(7-dimethylamino-4-methyl-3-coumarinyl)-maleimide (DACM). In frozen sections, dermal amyloids of skin-limited amyloidoses contained a large number of S-S bonds but no -SH groups [macular amyloidosis (9 cases), lichen amyloidosis ... More
Histidine-rich and cystine-rich keratohyalin granules in hamster cheek pouch epidermis. 1. Histochemical examination with a fluorescent thiol reagent (DACM).
AuthorsTezuka T
JournalCurr Probl Dermatol
PubMed ID6197248
'Hamster cheek pouch epidermis was examined with a thiol-specific fluorescent reagent [7-(N-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM)], with the Pauly reagent and by electron microscopy. There were many keratohyalin granules in the cells of the stratum granulosum where they were positively stained with the Pauly reagent, and were fluorescent as a whole, rather ... More
Effects of preparation and fixation on three quantitative fluorescent cytochemical procedures.
AuthorsCurtis SK, Cowden RR
JournalHistochemistry
PubMed ID6157658
'A series of experiments was undertaken in which cells dissociated from the abdominal lymph nodes of mice were lightly centrifuged into slides and fixed either wet or after drying in 70% ethanol, 1% glutaraldehyde, 1% formaldehyde, or neutral formalin. Three fluorescent cytochemical methods were evaluated: staining of DNA with mithramycin; ... More
Temperature-induced transitions of porcine intestinal brush border membranes.
AuthorsOhyashiki T, Takeuchi M, Kodera M, Mohri T
JournalBiochim Biophys Acta
PubMed ID7093271
'Thermotropic transitions of the membrane components in porcine intestinal brush border membranes were studied by means of fluorimetry using a fluorogenic thiol reagent, N-[7-dimethylamino-4-methylcoumarinyl]maleimide (DACM), and a lipophilic fluorescent probe, 1,6-diphenyl-1,3,5-hexatriene (DPH). 1. The reactivity of the sulfhydryl groups of the membrane proteins with DACM was dependent on temperature, with ... More
Variation of the distribution pattern of SH groups and SS linkages in the human epidermis.
AuthorsIto M, Suzuki M, Maruyama T, Sato Y, Hashimoto K
JournalJ Cutan Pathol
PubMed ID3771873
'Normal epidermis of various sites of human body was examined by N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide (DACM) method for SH groups and SS linkages. The horny cells in the palm and sole showed a gradual occurrence of a SH membranous pattern, while those in other sites revealed an abrupt formation of the SH membranous ... More
Fretting about FRET: correlation between kappa and R.
AuthorsVanBeek DB, Zwier MC, Shorb JM, Krueger BP
JournalBiophys J
PubMed ID17384068
'Molecular dynamics simulations were used to examine the structural dynamics of two fluorescent probes attached to a typical protein, hen egg-white lysozyme (HEWL). The donor probe (D) was attached via a succinimide group, consistent with the commonly-used maleimide conjugation chemistry, and the acceptor probe (A) was bound into the protein ... More
Electron microscopic visualization of the ATPase site of myosin by photoaffinity labeling with a biotinylated photoreactive ADP analog.
AuthorsSutoh K, Yamamoto K, Wakabayashi T
JournalProc Natl Acad Sci U S A
PubMed ID2934740
'An ADP analog carrying a biotin moiety and a photoreactive group was synthesized. In the presence of vanadate ion (Vi), the analog was tightly trapped into the ATPase site of heavy meromyosin (HMM) or myosin subfragment 1 (S1) in an ADP analog/ATPase site molar ratio of 1:1. UV illumination on ... More
Characterization of the reactivity of sulphydryl groups in tryptophanase by a dual-monitoring high-performance liquid chromatographic system with a site-directed fluorescent reagent.
AuthorsHonda T, Cacace MG, Sada A, Tokushige M
JournalJ Chromatogr
PubMed ID3558554
'Sulphydryl groups of E. coli tryptophanase (L-tryptophan indole lyase, E.C. 4.1.99.1) were made to react with a fluorescent maleimide derivative, N-(4-anilino-1-naphthyl)maleimide(ANM). By carefully controlling the reaction conditions it was possible to limit the extent of sulphydryl group modification. The modified enzyme was digested with (L-1-tosylamide-2-phenylethyl chloromethyl ketone)-trypsin. The fluorescent peptides ... More
Fluorometric studies on the light chains of skeletal muscle myosin. I. Effects of temperature, ionic strength, divalent metal ions, and nucleotides.
AuthorsYamamoto K, Sekine T
JournalJ Biochem (Tokyo)
PubMed ID914809
'Light chains of skeletal muscle myosin were studied through the reactivity of their SH groups with a fluorescent thiol reagent, N-(7-dimethylamino-4-methylcoumarinyl) maleimide (DACM). The experiments were carried out by reacting the reagent with myosin for a short time and measuring the amounts of reacted dye by fluorometry after separating light ... More
Determination of 2-mercaptopropionylglycine in plasma and urine by high-performance liquid chromatography.
AuthorsKågedal B, Carlsson M, Denneberg T
JournalJ Chromatogr
PubMed ID3760058
'Methods for quantitative analysis of total and non-protein-bound 2-mercaptopropionylglycine (2-MPG) in plasma, and total 2-MPG in urine, have been developed. By reduction of urine, plasma or deproteinized plasma samples with tributylphosphine, 2-MPG is liberated from its disulphides, and after clean-up of the sample, 2-MPG is derivatized with N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide (DACM). The ... More
Preparation and characterization of fluorescent 50S ribosomes. Specific labeling of ribosomal proteins L7/L12 and L10 of Escherichia coli.
AuthorsZantema A, Maassen JA, Kriek J, Möller W
JournalBiochemistry
PubMed ID6125208
'So that the topographic and dynamic properties of the L7/L12--L10 complex in the 50S ribosome of Escherichia coli could be studied, methods and reagents were developed in order to introduce fluorescent groups at specific positions of these proteins. In the case of L7/L12, this was done by attaching an aldehyde ... More
Sulfhydryls on frog skeletal muscle membrane participate in contraction.
AuthorsOba T, Yamaguchi M
JournalAm J Physiol
PubMed ID2240191
'To examine the molecular mechanism underlying contractile activation, we studied effects of a sulfhydryl reagent, N-(7-dimethylamino-4-methylcoumarinyl)maleimide (DACM), on twitch, Ag(+)-induced contraction, and K+ and caffeine contractures in single toe muscle fibers of frog. DACM suppressed twitch and Ag(+)-induced contraction, dose dependently, but not caffeine contracture. K+ contracture also was decreased ... More
Pathogenesis in pili torti: morphological study.
AuthorsMaruyama T, Toyoda M, Kanei A, Morohashi M
JournalJ Dermatol Sci
PubMed ID7528051
'To clarify the mechanism of pili torti formation, hair materials obtained from a patient with the typical classical type of pili torti were examined. Light microscopy, scanning electron microscopy (SEM), transmission electron microscopy, (TEM) and 3-D analysis revealed that: (1) The hair shaft was partly flattened. The hairs twisted clockwise ... More
Sulphhydryl and disulphide stainings of subepidermal hyaline bodies.
AuthorsDanno K, Horio T
JournalBr J Dermatol
PubMed ID6165371
'The localization of sulphhydryl (-SH) groups and disulphide (-S-S-) bonds in subepidermal hyaline bodies was examined using histochemical techniques with the specific fluorescent thiol reagent (DACM). In lichen planus, hyaline bodies showed -S-S- but not -SH staining, suggesting that erythematosus and dermatomyositis, there were not only -S-S-positive hyaline bodies, considered ... More
A continuous fluorescent assay for measuring protease activity using natural protein substrate.
AuthorsFarmer WH, Yuan ZY
JournalAnal Biochem
PubMed ID1785689
'A continuous caseinolytic activity assay has been developed and characterized with trypsin, a serine protease, and transin, a metalloproteinase. Beta-casein labeled with both N-(7-dimethylamino-4-methylcoumarinyl)-maleimide (DACM) and fluorescein isothiocyanate (FITC) is used as the substrate in this assay. The effect of proteolysis of the substrate is a reduction of the intermolecular ... More
Chemical modification of glycerinated stalks shows tyrosine residues essential for spasmoneme contraction of Vorticella sp.
AuthorsFang J, Zhang B, Chen N, Asai H
JournalZoolog Sci
PubMed ID15170056
'Chemical modification of glycerinated stalks of Vorticella with TNM is used to investigate the role of tyrosine residues in the Ca(2+)-induced contraction of the spasmoneme. Tetranitromethane (TNM) is often employed as a specific reagent for the nitration of tyrosine residues in a protein at neutral and slightly alkaline pHs although ... More
The histochemical demonstration of protein-bound sulfhydryl groups and disulfide bonds in human hair by a new staining method (DACM staining).
AuthorsTaneda A, Ogawa H, Hashimoto K
JournalJ Invest Dermatol
PubMed ID6159428
'A new fluorogenic maleimide DACM, i.e., N(7-dimethyl amino-4-methyl coumarinyl) maleimide, does not fluoresce by itself. It is specifically combined with --SH groups and becomes fluorescent (lambda ex: 400 nm, lambda em: 485 nm). At this range of excitation the frozen skin section has no native fluorescence because the emission maximum ... More
Hormonal effects on the sulfhydryl groups associated with intestinal brush border membrane proteins.
AuthorsTolosa de Talamoni N, Mykkanen H, Cai Q, Wasserman RH
JournalBiochim Biophys Acta
PubMed ID1654120
'Previous studies demonstrated that the administration of 1,25-dihydroxycholecalciferol (1,25(OH)2D3) to cholecalciferol-deficient chicks rapidly increases the reactivity and amount of the sulfhydryl (HS-) groups in intestinal brush border membranes (BBM). In the present study, the tissue and hormonal specificity of this effect was investigated. The HS- groups of intestinal and renal ... More
Thiols of myosin. VI. Effect of ADP on the reactivity of thiol groups other than S2.
AuthorsYamamoto K, Sekine T
JournalJ Biochem (Tokyo)
PubMed ID7358655
'Experiments were carried out to explore the location of thiol groups which became less reactive when S2 became more reactive in the presence of ADP. Myosin was first treated with N-ethylmaleimide to block S1 and then treated with a fluorogenic thiol reagent, N-(7-dimethyl-amino-4-methylcoumarinyl) maleimide (DACM), in the presence or absence ... More
The -SH and SS positive components in dermal connective tissues: a study by N-(7-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM) stain.
AuthorsKanamori M, Shimizu M
JournalArch Dermatol Res
PubMed ID3674965
'Although -SH and SS positive components in dermal connective tissue have been observed by N-(7-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM) stain, their nature is as yet unknown. In our study, we stained several organs such as human skin, rabbit aorta, auricle, and trachea using DACM stain, and studied the distribution of -SH groups ... More
Synthesis and application of two reagents for the introduction of sulfhydryl groups into proteins.
AuthorsMaassen JA, Thielen TP, Möller W
JournalEur J Biochem
PubMed ID6873065
'Two reagents are described which can be used for the introduction of sulfhydryl groups into proteins. Mercaptopropionylhydrazide modifies specifically periodate-oxidized N termini of proteins, provided that the N-terminal residue is serine or threonine. 3-(Phenyldithio)propionimidate introduces a disulfide bond at lysine residues of proteins. Reduction converts the disulfide into a sulfhydryl ... More
Pathogenesis in trichorrhexis invaginata (bamboo hair).
AuthorsIto M, Ito K, Hashimoto K
JournalJ Invest Dermatol
PubMed ID6547464
'The mechanism of formation of trichorrhexis invaginata, which is one of the characteristic hair anomalies seen in Netherton''s syndrome, was studied using plucked hairs obtained from 3 patients with the typical clinical features. The biopsy specimens used to examine the hair root were obtained from the scalp of one of ... More
Assignment of catalytically essential cysteine residues in aspartase by selective chemical modification with N-(7-dimethylamino-4-methylcoumarynyl)maleimide.
AuthorsIda N, Tokushige M
JournalJ Biochem (Tokyo)
PubMed ID3910645
'N-(7-Dimethylamino-4-methylcoumarynyl)maleimide (DACM), a fluorescent reagent for sulfhydryl groups, was employed to determine the functionally essential cysteine residues in aspartase from Escherichia coli. Analysis of the tryptic peptides containing DACM-labeled residues by reverse phase HPLC revealed that Cys-140 and Cys-430 were selectively modified, among 11 residues whose loci were recently determined ... More
New findings on the proteins of sebaceous glands.
AuthorsIto M, Suzuki M, Motoyoshi K, Maruyama T, Sato Y
JournalJ Invest Dermatol
PubMed ID6200547
'In order to understand the distribution and concentration of proteins with -SH groups or S-S linkages in sebaceous cells during differentiation and holocrine secretion of sebaceous glands, skin specimens from the inner side of ears of New Zealand white rabbits were examined histochemically and ultrastructurally. DACM (N-[7-dimethylamino-4-methyl-3-coumarinyl] maleimide) staining method ... More
Disulfide bond interchange in Escherichia coli-derived recombinant human interferon-beta 1 under denaturing conditions.
AuthorsKimura S, Utsumi J, Yamazaki S, Shimizu H
JournalJ Biochem (Tokyo)
PubMed ID3220830
'Disulfide bond interchange has been pointed out as a considerable problem in preparing recombinant proteins from Escherichia coli cells. This has been reported in the system of reducing denaturation followed by a refolding process, where incorrectly folded molecules are sometimes produced. As the possibility of disulfide bond interchange may also ... More
Use of N-(7-dimethylamino-4-methylcoumarinyl)maleinimide for the detection of cysteine-containing peptides in peptide maps.
AuthorsKlasen EC
JournalAnal Biochem
PubMed ID6179437
Fluorescent tracer method for protein SH groups. II.
AuthorsYamamoto K, Okamoto Y, Sekine T
JournalAnal Biochem
PubMed ID626373
Fluorescent thiol reagents. XII. Fluorescent tracer method for protein SH groups using N-(7-dimethylamino-4-methyl coumarinyl) maleimide. An application to the proteins separated by SDS-polyacrylamide gel electrophoresis.
AuthorsYamamoto K, Takamitsu S, Kanaoka Y
JournalAnal Biochem
PubMed ID869207
Demonstration of sulfhydryl and disulfide groups by a fluorescent maleimide procedure.
AuthorsCurtis SK, Cowden RR
JournalHistochemistry
PubMed ID7190963
Several fluorescent maleimide compounds were evaluated as possible substitutes for N-(4-aminophenyl)maleimide in the histochemical procedures developed by Sippel (1973, 1978a, b, 1980) for the demonstration of sulfhydryl and disulfide groups. The brightest and most selective fluorescence was obtained by using N-(7-dimethylamino-4-methylcoumarinyl)maleimide (DACM), although both eosin-5-maleimide and fluorescein-5-maleimide could also be ... More
Preparation of fluorescence-labeled and cross-linked subtilisin.
AuthorsTanizawa K, Mano T, Kanaoka Y
JournalChem Pharm Bull (Tokyo)
PubMed ID2186877
Streptomyces subtilisin inhibitor (SSI) is a protein characterized by both its potent inhibitory activity toward subtilisin and its structure, composed of two homologous subunits. It binds two molecules of subtilisin to form a tetrameric complex. Intermolecularly cross-linked subtilisin is expected to form a polymeric complex with SSI. This could provide ... More
SH abundant body in infantile digital fibromatosis--a histochemical study on distribution of sulfhydryl groups.
AuthorsKanamori M, Hashimoto K, Tanaka H, Shimizu M
JournalJ Dermatol
PubMed ID2177484
A case of infantile digital fibromatosis was studied with regard to the distribution of -SH groups and SS linkages, using N-(7-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM) stain. Tumor cells showed a strong fluorescence in the cytoplasm from the -SH stain. A brilliant -SH fluorescence was also homogenously present on the intracytoplasmic inclusions. SS ... More
Transition protein 1 from boar late spermatid nuclei having DNA-melting activity is a dimeric protein.
AuthorsAkama K, Sato H, Hasegawa S, Shimada I, Nakano M
JournalBiochem Mol Biol Int
PubMed ID9530514
Polyacrylamide gel electrophoretic behavior of boar transition protein 1, TP1, under dissociating and non-dissociating buffer conditions, and titration of fluorescently labeled TP1 with increasing amounts of TP1 showed that TP1 formed a dimer without intermolecular disulfide bond. TP1 dimer with intermolecular disulfide bond had similar DNA-melting activity to TP1, but ... More
Identification of sulfur groups in the outer plexiform layer of the newt retina.
AuthorsBall AK, Dickson DH
JournalExp Eye Res
PubMed ID6193987
The staining of newt photoreceptor synaptic vesicles with the zinc iodide-osmium (ZIO) method can be abolished by prior incubation in amphibian Ringer's containing 0.1 M-N-ethyl maleimide which specifically blocks -SH groups. The existence of high concentrations of sulfur groups in the outer plexiform layer of the newt retina has been ... More
Ouabain-binding site of (Na+ + K+)-ATPase in right-side-out vesicles has not an externally accessible SH group.
AuthorsScheiner-Bobis G, Zimmermann M, Kirch U, Schoner W
JournalEur J Biochem
PubMed ID3036511
The fluorescing sulfhydryl reagent N-(7-dimethylamino-4-methylcoumarinyl)maleimide (DACM) inactivates purified (Na+ + K+)-ATPase at 20 microM. This inactivation results in a decrease of the ouabain-binding capacity of the enzyme. Treatment of (Na+ + K+)-ATPase, embedded in right-side-out-oriented vesicles, by DACM does not affect ouabain binding to the enzyme. Incorporation of DACM into ... More
N-(7-dimethylamino-4-methylcoumarinyl)-maleimide (DACM): an alternative label for fluorescence tracing.
AuthorsNamihisa T, Saifuku K, Ishii H, Watanabe S, Sekine T
JournalJ Immunol Methods
PubMed ID6338112
N-(7-dimethylamino-4-methylcoumarinyl)maleimide (DACM), a fluorescent thiol reagent, has a high molar extinction coefficient and a high quantum yield when it reacts with protein sulphhydryl bonds. DACM proved to be effective as a fluorescent probe when used for labelling rabbit immunoglobulin G anti-human gamma-chain (IgG). DACM-labelled rabbit immunoglobulin G anti-human gamma-chain conjugates ... More
Fluorescent staining of microfilaments with heavy meromyosin labeled with N-(7-dimethylamino-4-methylcoumarinyl) maleimide.
AuthorsNamihisa T, Tamura K, Saifuku K, Imanari H, Kuroda H, Kanaoka Y, Okamoto Y, Sekine T
JournalJ Histochem Cytochem
PubMed ID6989895
For fluorescent staining of microfilaments in cells, heavy meromyosin (HMM) or subfragment-1 (S-1) was labeled with a novel thiol-directed fluorescent dye, N-(7-dimethylamino-4-methylcoumarinyl) maleimide (DACM), instead of the usual dyes, such as fluorescein-isothiocyanate (FITC). DACM-labeled HMM or S-1 gave characteristic fluorescence patterns to a variety of cell types similar to those ... More
Assessment of sulfhydryl group in individual rat lens protein subunits during galactose cataract development.
AuthorsPan S, Hua JC, Calvin HI, Fu SC
JournalYan Ke Xue Bao
PubMed ID7843379
A specific reagent DACM [N-(7-Dimethylamino-4-methyl-3-coumarinyl)maleimide] is used to study the -SH groups in lens proteins of normal and galactose cataractous rats. DACM when reacts readily with -SH groups form strong fluorescent adducts. The two -dimensional electrophoresis with DACM pre-labeled proteins is a simple and sensitive method for detecting -SH groups ... More
Monomaleimidogold labeling of the gamma subunit of the Escherichia coli F1 ATPase examined by cryoelectron microscopy.
AuthorsWilkens S, Capaldi RA
JournalArch Biochem Biophys
PubMed ID1444442
A novel approach for locating sites of interest in a protein complex has been developed using monomaleimidonanogold (MMN). The Escherichia coli F1 ATPase, when prepared without the delta subunit, contains only a single reactive cysteine on one of the three copies of the alpha subunit. This site was reacted with ... More
Use of hybridization for distance measurement by fluorescence energy transfer in oligomeric proteins: distance between two functional sites in aspartase.
AuthorsMurase S, Kawata Y, Yumoto N
JournalBiochem Biophys Res Commun
PubMed ID8216244
Our previous studies suggested that in a tetrameric enzyme aspartase, Cys-140 and Trp-430 are located at or near the catalytic and activator sites, respectively. To estimate the distance between these two sites, fluorescence energy transfer between a single tryptophan (Trp-430) and a fluorescent group specifically attached to Cys-140 has been ... More
An actin-binding site on the 20K fragment of myosin subfragment 1.
AuthorsSutoh K
JournalBiochemistry
PubMed ID7138830
Myosin subfragment 1 (S1) was covalently labeled with a fluorescent dye, N-[7-(dimethylamino)-4-methyl-3-coumarinyl]maleimide (DACM), and then digested by trypsin to cleave S1 heavy chain into fragments. The DACM-labeled and trypsin-treated S1 was complexed with F-actin and treated with a zero-length cross-linker, 1-ethyl-3[3-(dimethylamino)propyl] carbodiimide (EDC). The cross-linking reaction generated a covalently linked ... More
Assembly of clathrin molecules on liposome membranes: a possible event necessary for induction of membrane fusion.
AuthorsMaezawa S, Yoshimura T
JournalBiochem Biophys Res Commun
PubMed ID2124108
Below pH6, clathrin induces fusion of liposomes containing phosphatidylserine (PS) [Maezawa et al. (1989) Biochemistry 28, 1422-1428]. Under similar conditions clathrin forms self-aggregates, suggesting that the associated form of clathrin may be involved in the fusion process. For examination of this possibility, the extent of fluorescence energy transfer from N-(p-(2-benzimidazolyl)phenyl)maleimide ... More
Dyskeratotic process of hyperkeratosis lenticularis perstans (Flegel). The abnormal formation of cystine-rich protein.
AuthorsTezuka T
JournalDermatologica
PubMed ID7117641
The dyskeratotic changes in the lesions of hyperkeratosis lenticularis perstans were examined electron microscopically and histochemically. Under electron microscopy, the formation of keratohyalin granules was reduced and there was a lack of lamellar granule formation. In addition, the catabolic processes were disturbed, and ribosomal and mitochondrial structures remained, and lipid ... More
Topography of the yeast ATP synthase F0 sector by using cysteine substitution mutants. Cross-linkings between subunits 4, 6, and f.
The arrangement of the N-terminal part of subunit 4 (subunit b) has been studied by the use of mutants containing cysteine residues in a loop connecting the two N-terminal postulated membrane-spanning segments. Labelling of the mutated subunit 4 by the fluorescent probe N-(7-(dimethylamino)-4-methyl-3-coumarinyl)maleimide revealed that the sulfhydryl groups were modified ... More
Location of SH1 and SH2 along a heavy chain of myosin subfragment 1.
AuthorsSutoh K
JournalBiochemistry
PubMed ID7248283
Two reactive SH groups (SH1 and SH2) of myosin subfragment 1 (S-1) were selectively labeled with a fluorescent dye, N-[7-(dimethylamino)-4-methyl-3-coumarinyl]maleimide (DACM). When the DACM-S-1 was digested with trypsin, 95K heavy chain was cleaved into 50K, 25K, and 20K fragments, and the fluorescent labels incorporated into SH1 and SH2 were exclusively ... More
Manipulation of carrier proteins in antibiotic biosynthesis.
Engineering biosynthetic pathways into suitable host organisms has become an attractive venue for the design, evaluation, and production of small molecule therapeutics. Polyketide (PK) and nonribosomal peptide (NRP) synthases have been of particular interest due to their modular structure, yet routine cloning and expression of these enzymes remains challenging. Here ... More
Cysteinyl residues of Escherichia coli recA protein.
The Escherichia coli recA protein has three cysteinyl residues at positions 90, 116, and 129. All of them are reactive with 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB). In the presence of ATP or ADP, only one cysteinyl residue reacts with DTNB. The residue was also reactive with N-[7-(dimethylamino)-4-methylcoumarinyl]maleimide (DACM) in the presence of ... More
Quantitative determination of free thiol and disulphide groups by a fluorescent maleimide procedure.
AuthorsBroekaert D, Coucke P, Nsabumukunzi S, Reyniers P, van Oostveldt P, Kluyskens P, Gillis E
JournalActa Histochem
PubMed ID6179125
To determine the quantitative reliability of thiol histofluorometry, the distribution of protein thiol and disulphide groups was reinvestigated in normal human epidermis, labelled with DACM-3 after cryocutting. Microscopical observations roughly confirmed, that living keratinocytes and inner stratum corneum are the main sites of free thiol groups, while disulphide crosslinks are ... More
The fluroescent profiles of keratohyalin granules of newborn rat epidermis with a new fluorescent thiol reagent (DACM).
AuthorsTezuka T, Hirai R, Ogawa H
JournalActa Derm Venereol
PubMed ID82347
A highly specific fluorescent thiol reagent, N-(7-Dimethylamino-4-methyl-coumarinyl)maleimide (DACM) has been used to examine the newborn rat skin in order to elucidate the cystine-rich keratohyalin granules or dense homogeneous deposits. After reduction of the disulphide bond with 2-mercaptoethanol, the periphery of the stratum corneum cells showed a very distinct fluorescence and ... More
Propidium iodide and the thiol-specific reagent DACM as a dye pair for fluorescence resonance energy transfer analysis: an application to mouse sperm chromatin.
AuthorsBottiroli G, Croce AC, Pellicciari C, Ramponi R
JournalCytometry
PubMed ID8168398
The dyes N-(-7-dimethyl-amino-4-methyl-coumarinyl) maleimide and propidium iodide, specific for the thiol group and DNA, respectively, were considered as a donor-acceptor couple suitable for investigating "in situ" the relative spatial distribution of DNA and protamines in mouse spermatozoa chromatin. The two dyes are characterized by favourable spectral properties, so that a ... More
Oxidative lesion to membrane proteins in senescent erythrocytes.
AuthorsBrovelli A, Seppi C, Castellana AM, De Renzis MR, Blasina A, Balduini C
JournalBiomed Biochim Acta
PubMed ID1696812
The oxidative lesion undergone by membrane proteins in senescent human erythrocytes was evaluated by assaying their MetSO and thiol group content in ghosts and the amount of a coumarinyl derivative of maleimide, the DACM, bound by individual membrane proteins after treatment of erythrocytes of different age with this reagent. Quantitation ... More
Nucleus of the boar spermatozoon: structure and modifications in frozen, frozen-thawed, or sodium dodecyl sulfate-treated cells.
AuthorsCourtens JL, Paquignon M, Blaise F, Ekwall H, Plöen L
JournalMol Reprod Dev
PubMed ID2483517
After cryosubstitution and Epon embedding, or after Nanoplast embedding and very thin sectioning, the chromatin of ejaculated or diluted boar spermatozoa appears to be formed of DNA fibers embedded in a quite homogeneous matrix. After sodium dodecyl sulfate (SDS) treatment, and to a lesser extent after freeze-thawing, the DNA fibers ... More
Limited proteolysis of a chemically modified third component of human complement, C3, by cathepsin G of human leukocytes.
AuthorsMatsuda T, Nagasawa S, Koide T, Koyama J
JournalJ Biochem (Tokyo)
PubMed ID4044552
We have investigated the limited proteolysis of the third component of complement, C3, by a human leukocyte protease, cathepsin G, by using a chemically modified C3, which was prepared by treatment of C3 with methylamine and a fluorescent thiol reagent, N-(dimethylamino-4-methylcoumarinyl)-maleimide (DACM) and was thus named DACM-C3me. Although native C3 ... More
Enzyme cytochemical techniques for metabolic mapping in living cells, with special reference to proteolysis.
AuthorsBoonacker E, Van Noorden CJ
JournalJ Histochem Cytochem
PubMed ID11724895
Specific enzymes play key roles in many pathophysiological processes and therefore are targets for therapeutic strategies. The activity of most enzymes is largely determined by many factors at the post-translational level. Therefore, it is essential to study the activity of target enzymes in living cells and tissues in a quantitative ... More
Evidence for membrane protein oxidation during in vivo aging of human erythrocytes.
Oxidative lesions to membrane proteins were studied in human erythrocytes of different age and were evaluated on ghost membrane preparations by assaying thiol and methionine sulphoxide groups, and in situ on intact cells, after treating erythrocytes with the fluorochrome N-(7-dimethyl-amino-4-methyl-coumarinyl) maleimide (DACM). DACM reacts with thiol groups and the amount ... More
Changes in fluorescence energy transfer between sulfhydryl fluorescent residues during ouabain sensitive Na+,K+-ATP hydrolysis.
AuthorsSakuraya M, Taniguchi K, Suzuki K, Kudo A, Nakamura S, Iida S
JournalJpn J Pharmacol
PubMed ID2821309
Na+,K+-ATPase from pig kidney was sequentially modified with two different sulfhydryl fluorescent reagents, N-[p-(2-benzimidazolyl)phenyl]maleimide (BIPM) and N-[7-dimethylamino 4-coumarinyl]maleimide (DACM). The preparation thus obtained contained 3 and 2 moles of each residue in the alpha-chain. When the BIPM residues were excited at 313 nm, ouabain sensitive decrease and increase in the ... More
Conformational change of the foot protein of sarcoplasmic reticulum as an initial event of calcium release.
AuthorsOhkusa T, Kang JJ, Morii M, Ikemoto N
JournalJ Biochem (Tokyo)
PubMed ID1869514
Heavy sarcoplasmic reticulum vesicles were labeled with the thiol-reacting fluorescent probe N-(7-dimethylamino-4-methyl-4-coumarinyl)maleimide (DACM), and the DACM-labeled foot protein moiety was purified. The fluorescence intensity of the DACM attached to the foot protein decreased by the addition of low (activating) concentrations of ryanodine, while it increased at higher (inhibitory) concentrations, suggesting ... More
Reactivity of sulfhydryl groups in the brush-border membranes of chick duodena is increased by 1,25-dihydroxycholecalciferol.
AuthorsMykkanen HM, Wasserman RH
JournalBiochim Biophys Acta
PubMed ID2317503
The total amount and reactivity of SH-groups were determined in isolated duodenal brush-border membranes from rachitic chicks given 1,25-dihydroxyvitamin D-3 (1,25(OH)2D3) before isolation of the membranes. 1,25(OH)2D3 treatment significantly increased the total amount of SH groups (9.7 +/- 2.3 vs. 23.9 +/- 2.1, P less than 0.001, n = 6) ... More
Fluorescence conformational probe study of calcium release from sarcoplasmic reticulum.
AuthorsMorii M, Danko S, Kim DH, Ikemoto N
JournalJ Biol Chem
PubMed ID2418023
Sarcoplasmic reticulum isolated from rabbit skeletal muscle was labeled with a limited (0.625 nmol/mg sarcoplasmic reticulum protein) amount of the fluorescent thiol reagent N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide (DACM). The fluorescence intensity of the membrane-attached DACM decreased concurrently with (Ca2+ and caffeine)-induced Ca2+ release, depolarization-induced Ca2+ release and Ca2+-dependent dependent passive efflux of Ca2+. ... More
Evaluation of methods for the quantitation of cysteines in proteins.
AuthorsWright SK, Viola RE
JournalAnal Biochem
PubMed ID9866701
Several methods for the quantitation of cysteines in proteins have been evaluated and compared. Titration of protein sulfhydryl groups with 5,5'-dithiobis(2-nitrobenzoate) (DTNB) under carefully controlled conditions has extended the detection limits of this method with high accuracy and reproducibility. Results are reported for a variety of enzymes containing a range ... More
Increased SS bonds in chronic solar elastosis: a study with N-(7-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM) stain.
AuthorsPravatà G, Noto G, Aricò M
JournalJ Dermatol Sci
PubMed ID8193080
We have studied the distribution of SH groups and SS linkages in solar elastosis, in comparison with that in aged and juvenile sun-unexposed skin, using N-(7-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM) stain. In heavy solar elastosis we found increased fluorescence for SS bonds in the upper and middle reticular dermis in the elastotic ... More
Fluorescence and electron microscopic study of intracellular F-actin in concanavalin A-treated and cytochalasin B-treated Ehrlich ascites tumor cells.
AuthorsWatanabe S, Takehara Y, Fujii Y, Okimasu E, Moromizato Y, Sasaki J
JournalActa Med Okayama
PubMed ID3825593
To investigate the involvement of actin filaments in concanavalin A (Con A)-induced cap formation and cytochalasin B (CB)-induced zeiotic knob migration, the distribution of F-actin was studied in Con A-treated and CB-treated Ehrlich ascites tumor cells (EATC) by fluorescence microscopy using heavy meromyosin conjugated with a fluorescent dye, N-(7-dimethylamino-4-methylcoumarinyl) maleimide, ... More
Energy transfer between fluorescent dyes attached to Ca2+,Mg2+-ATPase in the sarcoplasmic reticulum.
AuthorsYantorno RE, Yamamoto T, Tonomura Y
JournalJ Biochem (Tokyo)
PubMed ID6140262
Sarcoplasmic reticulum (SR) isolated from rabbit skeletal muscle was solubilized with a nonionic detergent, dodecyl octaethyleneglycol monoether (C12E8), at a weight ratio of detergent to protein of greater than 10, so that the Ca2+, Mg2+ dependent ATPase existed mainly in a monomeric form (7). The solubilized ATPase was reacted with ... More
Studies on conformational transitions of Ca2+,Mg2+-adenosine triphosphatase of sarcoplasmic reticulum. II. Conformational characteristics of stabilized reaction intermediates as revealed by fluorescent and paramagnetic probes.
AuthorsYasuoka-Yabe K, Tsuji A, Kawakita M
JournalJ Biochem (Tokyo)
PubMed ID6139371
Various reaction intermediates of sarcoplasmic reticulum Ca2+,Mg2+-ATPase were stabilized and accumulated by modifying a specific SH group or by using nucleotide analogs. Conformational changes of the Ca2+,Mg2+-ATPase during the catalytic cycle were studied in the stabilized intermediates by the use of fluorescent and spin probes, which were introduced at specific ... More
Modification of subunit b of the F0 complex from Escherichia coli ATP synthase by a hydrophobic maleimide and its effects on F0 functions.
AuthorsSchneider E, Altendorf K
JournalEur J Biochem
PubMed ID2866095
Purified F0 from Escherichia coli ATP synthase was labelled with N-(7-dimethylamino-4-methyl-coumarinyl)-maleimide (DACM), a hydrophobic reagent which forms a stable, strongly fluorescent adduct with SH groups. Sodium dodecyl sulfate gel electrophoresis clearly demonstrated that subunit b was exclusively labelled, most likely at Cys-21, the only cysteine residue in E. coli F0. ... More
Enhancement of sulfhydryl group availability in the intestinal brush border membrane by deficiencies of dietary calcium and phosphorus in chicks.
Authorsde Talamoni NT, Mykkanen H, Wasserman RH
JournalJ Nutr
PubMed ID2213248
The reactivity and availability of sulfhydryl (-SH) groups in brush border membranes (BBM) from chicks adapted to a calcium-deficient (low Ca) or a phosphorus-deficient (low P) diet were determined. The calbindin-D28K concentrations of the intestinal mucosa of the low Ca and low P groups were both increased approximately 2.5-fold, demonstrating ... More
Localization of virginiamycin S binding site on bacterial ribosome by fluorescence energy transfer.
AuthorsDi Giambattista M, Thielen AP, Maassen JA, Möller W, Cocito C
JournalBiochemistry
PubMed ID3087416
Virginiamycin S, a type B synergimycin inhibiting protein synthesis in bacteria, competes with erythromycin for binding to the 50S ribosomal subunits; the mechanism of action of the two antibiotics is unclear. Energy-transfer experiments between virginiamycin S (which is endowed with inherent fluorescence due to its hydroxypicolinyl moiety) and fluorescent coumarinyl ... More
Effects of tetraphenylboron-induced increase in inner surface charge on Ca2+ release from sarcoplasmic reticulum.
AuthorsLiu GH, Oba T
JournalJpn J Physiol
PubMed ID2086992
We studied the relationship between surface charge and release of Ca2+ in the heavy sarcoplasmic reticulum (SR) of skeletal muscle. The inner and outer surface potentials and charge densities of the membrane treated with a lipophilic anion, tetraphenylboron (TPB-), were measured using 1-anilino-8-naphthalene-sulfonate fluorescence. Ca2+ was loaded passively or actively ... More
Increase of the molecular rigidity of the protein conformation in the intestinal brush-border membranes by lipid peroxidation.
AuthorsOhyashiki T, Ohtsuka T, Mohri T
JournalBiochim Biophys Acta
PubMed ID3355823
The effect of lipid peroxidation on the protein conformation of the porcine intestinal brush-border membranes was studied using a fluorogenic thiol reagent, N-[7-dimethylamino-4-methylcoumarinyl]maleimide (DACM). By a kinetic analysis of the reaction of the membranes with DACM, it was shown that the reaction rate of the SH groups (SHf) of the ... More
Investigation of the role of cysteines in catalysis by prostaglandin endoperoxide synthase.
AuthorsKennedy TA, Smith CJ, Marnett LJ
JournalJ Biol Chem
PubMed ID7961646
The importance of cysteine residues in the cyclooxygenase activity of prostaglandin endoperoxide synthase (PGHS) was investigated using cysteine-specific reagents and site-directed mutagenesis. N-(7-Dimethyl-amino-4-methyl-3-coumarinyl)maleimide (DACM), a hydrophobic maleimide, inactivated both cyclooxygenase and peroxidase activities of apoPGHS in a time-dependent manner but did not affect holoPGHS. Heme titration experiments indicated that modification ... More
Accumulation of fluorescently labeled actin in the cortical layer in sea urchin eggs after fertilization.
AuthorsHamaguchi Y, Mabuchi I
JournalCell Motil Cytoskeleton
PubMed ID3359492
Actin from sea urchin eggs was fluorescently labeled with fluorescein isothiocyanate (FITC), N-(7-dimethylamino-4-methylcoumarinyl)-maleimide (DACM), or 5-iodoacetamidofluorescein (IAF) and microinjected into sea urchin eggs and oocytes. It distributed evenly in the cytoplasm of unfertilized eggs. Upon fertilization, actin accumulated first around the sperm binding site and, soon afterwards, in the fertilization ... More
Sulfhydryl oxidase (SOx) from mouse epidermis: molecular cloning, nucleotide sequence, and expression of recombinant protein in the cultured cells.
AuthorsMatsuba S, Suga Y, Ishidoh K, Hashimoto Y, Takamori K, Kominami E, Wilhelm B, Seitz J, Ogawa H
JournalJ Dermatol Sci
PubMed ID12354420
Skin sulfhydryl oxidase (SOx) is an enzyme that catalyzes disulfide (S-S) cross-linking through the oxidation of sulfhydryl compounds in the skin. In this study, using the enzyme purified from rat seminal vesicle, we obtained peptide sequences for SOx by mass spectrometry. We then searched for SOx nucleotides corresponding highly to ... More
Chemical evidence for the existence of activated G-actin.
AuthorsShu WP, Wang D, Stracher A
JournalBiochem J
PubMed ID1575699
Globular actin (G-actin) will polymerize to form filamentous actin (F-actin) under physiological ionic conditions, and is known to be regulated by univalent and bivalent cations, such as K+ and Mg2+. The current concept of this process involves four steps: activation, nucleation, elongation and annealing. Evidence for the existence of activated ... More
Trypanosoma cruzi phospho enol pyruvate carboxykinase (ATP-dependent): transition metal ion requirement for activity and sulfhydryl group reactivity.
AuthorsJurado LA, Machín I, Urbina JA
JournalBiochim Biophys Acta
PubMed ID8547343
We studied the transition metal ion requirements for activity and sulfhydryl group reactivity in phospho enol pyruvate carboxykinase (PEP-carboxykinase; ATP:oxaloacetate carboxylase (transphosphorylating), EC 4.1.1.49), a key enzyme in the energy metabolism of the protozan parasite Trypanosoma (Schizotrypanum) cruzi. As for other PEP-carboxykinases this enzyme has a strict requirement of transition ... More
The effects of ionic strength on the protein conformation and the fluidity of porcine intestinal brush border membranes. Fluorometric studies using N-[7-dimethylamino-4-methylcoumarinyl]maleimide and pyrene.
AuthorsOhyashiki T, Taka M, Mohri T
JournalJ Biol Chem
PubMed ID3997850
The effects of ionic strength on the conformation around the SH groups of the proteins and the lipid fluidity of porcine intestinal brush border membranes were studied using two fluorescent dyes, N-[7-dimethylamino-4-methylcoumarinyl]maleimide (DACM) and pyrene. The extent of DACM labeling to the SH groups of the membrane proteins was accelerated ... More
Studies on conformational transitions of Ca2+, Mg2+-adenosine triphosphatase of sarcoplasmic reticulum. I. Selective labeling of functionally distinct sulfhydryl groups with conformational probes and evidence for a Ca2+-dependent conformational change.
AuthorsYasuoka-Yabe K, Kawakita M
JournalJ Biochem (Tokyo)
PubMed ID6139370
Several maleimide derivatives of potential usefulness as conformational probes were tested for reactivity toward SH groups of Ca2+, Mg2+-ATPase of sarcoplasmic reticulum. These include three fluorescent labels, N-(1-anilinonaphthyl-4)maleimide (ANM), N-(p-(2-benzimidazolyl)phenyl)maleimide (BIPM), and N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide (DACM), and a spin label, 4-maleimido-2,2,6,6-tetramethylpiperidinooxyl (MSL). These reagents also exhibit a selective reactivity toward SH groups ... More
The functions of the ninth component of human complement are sustained by disulfide bonds with different susceptibilities to reduction.
AuthorsHatanaka M, Seya T, Inai S, Shimizu A
JournalBiochim Biophys Acta
PubMed ID7947973
Purified C9 with expected hemolytic and polymerizing activities was found to contain approximately 0.2 mol of sulfhydryl groups/mol of C9. By proteolysis of C9 with labeled SH groups, the SH residues on intact C9 were mapped to Cys-359 and Cys-384 which, presumably, form an intra-domain disulfide bond in the intact ... More
The substrate of epidermal proteases--effects of in situ activation of epidermal thiol proteases on stratum granulosum cells.
AuthorsTakahashi M, Tezuka T
JournalJ Dermatol
PubMed ID2656803
In newborn rat skin, disulfide bonds exist in the cell membrane of the stratum corneum and in the cytoplasm of the stratum granulosum. However, in the latter region, they can be detected after ethanol fixed epidermis remains for 2 weeks at 4 degrees C. Thus, the idea arose that the ... More
GroEL locked in a closed conformation by an interdomain cross-link can bind ATP and polypeptide but cannot process further reaction steps.
AuthorsMurai N, Makino Y, Yoshida M
JournalJ Biol Chem
PubMed ID8910440
It has been believed that when GroEL binds to GroES its apical domain moves upward and outward. To inhibit this "opening" movement, its equatorial and apical domains were cross-linked through a disulfide bond between mutationally introduced cysteine residues at the positions of Asp-83 and Lys-327. To avoid possible undesired cross-linking, ... More
Postulated role of calsequestrin in the regulation of calcium release from sarcoplasmic reticulum.
AuthorsIkemoto N, Ronjat M, Mészáros LG, Koshita M
JournalBiochemistry
PubMed ID2790030
Ca2+ release from heavy sarcoplasmic reticulum (SR) vesicles was induced by 2 mM caffeine, and the amount (A) and the rate constant (k) of Ca2+ release were investigated as a function of the extent of Ca2+ loading. Under both passive and active loading conditions, the A value increased monotonically in ... More
Changes in SH reactivity of the protein in porcine intestinal brush-border membranes associated with lipid peroxidation.
AuthorsOhyashiki T, Sakata N, Matsui K
JournalJ Biochem (Tokyo)
PubMed ID8206871
The effects of lipid peroxidation on the SH reactivity of the proteins in porcine intestinal brush-border membranes were examined using a fluorogenic thiol reagent, N-[7-dimethyl-amino-4-methylcoumarinyl]maleimide (DACM) in relation to lipid organization. Changes in the lipid organization were assessed by measurement of the rate of incorporation of 1,6-diphenyl-1,3,5-hexatriene (DPH) into the ... More
Chemical modifications of chicken liver pyruvate carboxylase: evidence for essential cysteine-lysine pairs and a reactive sulfhydryl group.
AuthorsWerneburg BG, Ash DE
JournalArch Biochem Biophys
PubMed ID8512310
Inactivations of chicken liver pyruvate carboxylase with N-(7-dimethylamino-4-methyl-3-coumarinyl)maleimide (DACM) and o-phthalaldehyde (o-PA) have identified cysteine and lysine residues that are essential for catalytic activity. Protection experiments suggest that the modified residues are located in or near the first and second subsites. At a one- to two-fold molar excess over active ... More