Fluorescence resonance energy transfer between sites in G-actin. The spatial relationship between Cys-10, Tyr-69, Cys-374, the high-affinity metal and the nucleotide.
AuthorsBarden JA, dos Remedios CG
JournalEur J Biochem
PubMed ID3665911
'Intramonomer fluorescence resonance energy transfer spectroscopy was employed to investigate the spatial relationship between labels attached to the residues Cys-10, Tyr-69, Cys-374, the high-affinity metal binding site and the nucleotide binding site in G-actin. The separation between the fluorescence donor 5-(dimethylamino)naphthalene-1-sulphonyl (Dns) chloride (dansyl chloride) used to label Tyr-69 and ... More
Regulation of the rate and extent of phospholipase C beta 2 effector activation by the beta gamma subunits of heterotrimeric G proteins.
AuthorsRunnels LW, Scarlata SF
JournalBiochemistry
PubMed ID9799521
'The activity of mammalian phosphoinositide-specific phospholipase C beta 2 (PLC-beta 2) is regulated by the alpha q family of G proteins and by beta gamma subunits. We measured the affinity between the laterally associating PLC-beta 2 and G beta gamma on membrane surfaces by fluorescence resonance energy transfer. Using a ... More
Relocation of Cys374 of actin induced by labeling with fluorescent dyes.
AuthorsYasunaga T, Wakabayashi T
JournalJ Biochem (Tokyo)
PubMed ID11173519
'Cys374 is an important landmark of actin, because its sulfhydryl group is reactive and can be labeled with various reagents. The atomic coordinates of actin Cys374 have been determined by X-ray crystallography of co-crystals of actin with either profilin or gelsolin. However, the positions of Cys374 in the crystals determined ... More
Interhead fluorescence energy transfer between probes attached to translationally equivalent sites on the regulatory light chains of scallop myosin.
AuthorsChantler PD, Tao T
JournalJ Mol Biol
PubMed ID3820308
'Interhead fluorescence energy transfer studies between probes located at translationally equivalent sites on the two heads of scallop myosin indicates that the distance between such sites is no less than 50 A. Regulatory light chains, possessing either one (Mercenaria, chicken gizzard) or two (Loligo, rabbit skeletal) sulfhydryl groups, were modified ... More
The rates of switching movement of troponin T between three states of skeletal muscle thin filaments determined by fluorescence resonance energy transfer.
AuthorsShitaka Y, Kimura C, Miki M
JournalJ Biol Chem
PubMed ID15548522
'Troponin (Tn) plays the key roles in the regulation of striated muscle contraction. Tn consists of three subunits (TnT, TnC, and TnI). In combination with the stopped-flow method, fluorescence resonance energy transfer between probes attached to Cys-60 or Cys-250 of TnT and Cys-374 of actin was measured to determine the ... More
Resolution of a signal transfer region from a general binding domain in gbeta for stimulation of phospholipase C-beta2.
AuthorsBuck E, Li J, Chen Y, Weng G, Scarlata S, Iyengar R
JournalScience
PubMed ID10037604
'Signaling by guanine nucleotide-binding proteins (G proteins) involves sequential protein-protein interactions. G protein-betagamma subunit (Gbetagamma) interactions with phospholipase C-beta2 (PLC-beta2) were studied to determine if all Gbeta contacts are required for signaling. A peptide encoding Gbeta amino acid residues 86 to 105 stimulated PLC-beta2. Six residues (96 to 101) within ... More
Localization of the phalloidin and nucleotide-binding sites on actin.
AuthorsBarden JA, Miki M, Hambly BD, Dos Remedios CG
JournalEur J Biochem
PubMed ID3830158
'Phalloidin was found to block nucleotide exchange in F-actin, without interfering with nucleotide hydrolysis. This inhibition of nucleotide exchange occurs under conditions in which monomers are able to exchange. The distance separating a fluorescent chromophore attached to phalloidin from the nucleotide on actin was determined using fluorescence resonance energy-transfer spectroscopy. ... More
Determination of the affinities between heterotrimeric G protein subunits and their phospholipase C-beta effectors.
AuthorsRunnels LW, Scarlata SF
JournalBiochemistry
PubMed ID9931014
'Phosphatidylinositide-specific phospholipase C-betas play a key role in Ca2+ signaling and are specifically activated by the alphaq family of heterotrimeric G proteins and as well as betagamma subunits. We have determined the affinity between Gbetagamma subunits and GTPgammaS and GDP-liganded Galphaq subunits on membrane surfaces, and their respective affinities to ... More
Intradomain distances in the regulatory domain of the myosin head in prepower and postpower stroke states: fluorescence energy transfer.
AuthorsPalm T, Sale K, Brown L, Li H, Hambly B, Fajer PG
JournalBiochemistry
PubMed ID10529172
'The relative movement of the catalytic and regulatory domains of the myosin head (S1) is likely to be the force generating conformational change in the energy transduction of muscle [Rayment, I., Holden, H. M., Whittaker, M., Yohn, C. B., Lorenz, M., Holmes, K. C., and Milligan, R. A. (1993) Science ... More
Calcium-induced movement of troponin-I relative to actin in skeletal muscle thin filaments.
AuthorsTao T, Gong BJ, Leavis PC
JournalScience
PubMed ID2138356
'The role of troponin-I (the inhibitory subunit of troponin) in the regulation by Ca2+ of skeletal muscle contraction was investigated with resonance energy transfer and photo cross-linking techniques. The effect of Ca2+ on the proximity of troponin-I to actin in reconstituted rabbit skeletal thin filaments was determined. The distance between ... More
Resonance energy transfer between the active sites of rabbit muscle creatine kinase: analysis by steady-state and time-resolved fluorescence.
AuthorsGrossman SH
JournalBiochemistry
PubMed ID2765518
'Resonance energy transfer between the reactive thiols of rabbit muscle creatine kinase was evaluated. The reactive thiols are located at the active site, one occurring on each subunit of the dimeric protein that is known to be a constituent of the M-line structure of the myofibril. Transfer efficiency was evaluated ... More
Fluorescence resonance energy transfer between the nucleotide binding site and Cys-10 in G-actin and F-actin.
AuthorsMiki M, Barden JA, dos Remedios CG
JournalBiochim Biophys Acta
PubMed ID3089284
'Intramonomer fluorescence resonance energy transfer between the donor epsilon-ATP bound to the nucleotide site and the acceptor N-(4-dimethylamino-3,5-dinitrophenyl)maleimide (DDPM) or 4-dimethylaminophenyl-azophenyl-4''-maleimide bound to Cys-10 in G-actin was measured. The donor-acceptor distance was calculated to be about 40 A. The intermonomer energy transfer in F-actin occurring between epsilon-ADP and DABMI was ... More
Phosphorylation of smooth muscle myosin heads regulates the head-induced movement of tropomyosin.
AuthorsGraceffa P
JournalJ Biol Chem
PubMed ID10748060
'It has been shown that skeletal and smooth muscle myosin heads binding to actin results in the movement of smooth muscle tropomyosin, as revealed by a change in fluorescence resonance energy transfer between a fluorescence donor on tropomyosin and an acceptor on actin (Graceffa, P. (1999) Biochemistry 38, 11984-11992). In ... More
Regulatory properties of recombinant tropomyosins containing 5-hydroxytryptophan: Ca2+-binding to troponin results in a conformational change in a region of tropomyosin outside the troponin binding site.
AuthorsFarah CS, Reinach FC
JournalBiochemistry
PubMed ID10441151
'We have introduced tryptophan codons at different positions of the chicken alpha-tropomyosin cDNA (Monteiro, P. B., Lataro, R. C., Ferro, J. A., and Reinach, F. C. (1994) J. Biol. Chem. 269, 10461-10466) and employed a trp auxotrophic Escherichia coli strain to express the proteins in media containing either normal tryptophan, ... More
Excitation energy transfer studies of the proximity between tropomyosin and actin in reconstituted skeletal muscle thin filaments.
AuthorsTao T, Lamkin M, Lehrer SS
JournalBiochemistry
PubMed ID6224509
Structure of actin observed by fluorescence resonance energy transfer spectroscopy.
AuthorsMiki M, O'Donoghue SI, Dos Remedios CG
JournalJ Muscle Res Cell Motil
PubMed ID1534564
Fluorescence energy transfer between Cys-10 residues in F-actin filaments.
AuthorsMiki M, Barden JA, Hambly BD, dos Remedios CG
JournalBiochem Int
PubMed ID3089224
Fluorescence energy transfer was measured between Cys-10 residues in an F-actin filament using 5-[2-((iodoacetyl)amino)-ethyl]aminonaphthalene-1-sulphonic acid (1,5-IAEDANS) as a fluorescence energy donor and 4-dimethylaminophenylazophenyl-4'-maleimide (DABMI) as the acceptor. Both labels were covalently attached to Cys-10 residues in an F-actin filament. Taking the helical structure of the F-actin filament into consideration, the ... More
Recognition of cysteine-containing peptides through prompt fragmentation of the 4-dimethylaminophenylazophenyl-4'-maleimide derivative during analysis by MALDI-MS.
AuthorsBorges CR, Watson JT,
JournalProtein Sci
PubMed ID12824503
Under specified UV-MALDI conditions, the 4-dimethylaminophenylazophenyl-4'-maleimide (DABMI) derivative of cysteine-containing peptides undergoes prompt fragmentation to produce a characteristic mass spectral pattern. As reported previously by others, derivatization with chromophoric DABMI allows cysteine-containing peptides and proteins to be tracked during HPLC by absorbance at upper UV and visible wavelengths. Here, we ... More
pH-dependent structural transition in rabbit skeletal troponin C.
AuthorsWang CL, Zhan Q, Tao T, Gergely J
JournalJ Biol Chem
PubMed ID3597429
Although the crystal structure of troponin C is known (Herzberg, O., and James, M. N. G. (1985) Nature 313, 653-659; Sundaralingam, M., Bergstrom, R., Strasburg, G., Rao, S. T., Roychowdhury, P., Greaser, M., and Wang, B. C. (1985) Science 227, 945-948), its structure in solution, particularly under physiological conditions, has ... More
Investigations of the active site of Saccharomyces cerevisiae dolichyl-phosphate-mannose synthase using fluorescent labeled dolichyl-phosphate derivatives.
Dolichol-phosphate mannose (Dol-P-Man) is a key mannosyl donor for the biosynthesis of N-linked oligosaccharides as well as for O-linked oligosaccharides on yeast glycoproteins, and for the synthesis of the glycosyl-phosphatidylinositol anchor found on many cell surface glycoproteins. It is synthesized by Dol-P-Man synthase which is the only glycosyltransferase in the ... More
Proximity relationships between sites on myosin and actin. Resonance energy transfer determination of the following distances, using a hybrid myosin: those between Cys-55 on the Mercenaria regulatory light chain, SH-1 on the Aequipecten myosin heavy chain, and Cys-374 of actin.
AuthorsPark HS, Tao T, Chantler PD
JournalBiochemistry
PubMed ID2009259
Resonance energy transfer measurements have been made on hybrid myosins in order to map distances between sites on the regulatory light chain, heavy chain, and actin as well as to assess potential conformational changes of functional importance. Using scallop (Aequipecten) myosin hybrid molecules possessing clam (Mercenaria) regulatory light chains, we ... More
Fluorescence resonance energy transfer measurements of distances in actin and myosin. A critical evaluation.
Authorsdos Remedios CG, Miki M, Barden JA
JournalJ Muscle Res Cell Motil
PubMed ID3298315
The contractile proteins actin and myosin are of considerable biological interest. They are essential for muscle contraction and in eukaryotic cells they play a crucial role in most contractile phenomena. Over the years since the first fluorescence resonance energy transfer (FRET) paper appeared, an extensive body of literature has accumulated ... More
Fluorescence energy transfer studies on lima bean lectin. Distance between the subunit hydrophobic binding site and the thiol group essential for carbohydrate binding.
AuthorsKella NK, Roberts DD, Shafer JA, Goldstein IJ
JournalJ Biol Chem
PubMed ID6715322
Measurements of the efficiency of singlet-singlet energy transfer were used to determine the distance between the hydrophobic binding site and the thiol group required for carbohydrate-binding activity of lima bean lectin. 1-Anilino-8-naphthalenesulfonate, bound to the hydrophobic binding site by noncovalent interactions, was used as the donor. Two different nonfluorescent probes ... More
Distance measurements between metal-binding sites of calmodulin and from these sites to Cys-133 of troponin I in the binary complex.
AuthorsWang CL
JournalJ Biol Chem
PubMed ID3733748
Distances between the four Ca2+-binding sites of calmodulin (CaM) have been measured by fluorescence energy transfer techniques using Eu3+ and Tb3+ as energy donors and a number of other lanthanide ions (Ln3+) as acceptors. It was shown previously that lanthanide ions preferentially bind to sites I and II of CaM ... More
Movement of smooth muscle tropomyosin by myosin heads.
AuthorsGraceffa P
JournalBiochemistry
PubMed ID10508401
It has been proposed that during the activation of muscle contraction the initial binding of myosin heads to the actin thin filament contributes to switching on the thin filament and that this might involve the movement of actin-bound tropomyosin. The movement of smooth muscle tropomyosin on actin was investigated in ... More
High-affinity actin-binding nebulin fragments influence the actoS1 complex.
AuthorsRoot DD, Wang K
JournalBiochemistry
PubMed ID11170442
Human nebulin fragments, NA3 and NA4, corresponding to individual superrepeats display high-affinity interactions with individual actin protomers in cosedimentation and solid-phase binding assays. Stoichiometric analysis of nebulin fragment-induced actin polymerization and inhibition of actin-activated S1 ATPase indicate that one superrepeat influences multiple actin molecules along the F-actin filament, consistent with ... More
Spatial orientation and dynamics of the U1A proteins in the U1A-UTR complex.
AuthorsClerte C, Hall KB
JournalBiochemistry
PubMed ID10852733
The human U1A protein contains three distinct domains: the N-terminal RBD1 (amino acids 1-101), the C-terminal RBD2 (amino acids 195-282), and the linker region (amino acids 102-194). The RBD1 domains of two U1A proteins bind specifically to two internal loops in the 3' untranslated region (3' UTR) of its own ... More
Determination of the radial coordinate of Cys-374 in F-actin using fluorescence resonance energy transfer spectroscopy: effect of phalloidin on polymer assembly.
AuthorsMoens PD, Yee DJ, dos Remedios CG
JournalBiochemistry
PubMed ID7947715
In helically symmetric protein assemblies, fluorescence resonance energy transfer (FRET) spectroscopy can be used to determine the radial coordinates of fluorescent probes attached to specific amino acid side chains. This is done by separately labeling monomers with donor and acceptor probes, mixing them in different proportions, allowing the mixtures to ... More
Time-resolved fluorescence resonance energy transfer shows that the bacterial multidrug ABC half-transporter BmrA functions as a homodimer.
AuthorsDalmas O, Do Cao MA, Lugo MR, Sharom FJ, Di Pietro A, Jault JM
JournalBiochemistry
PubMed ID15766260
Members of the ATP-binding cassette (ABC) transporters share the same basic architecture, with a four-core domain made of two transmembrane plus two nucleotide-binding domains. However, a supramolecular organization has been detected in some ABC transporters, which might be relevant to physiological regulation of substrate transport. Here, the oligomerization status of ... More
Inhibitory region of troponin I: Ca(2+)-dependent structural and environmental changes in the troponin-tropomyosin complex and in reconstituted thin filaments.
AuthorsKobayashi T, Kobayashi M, Gryczynski Z, Lakowicz JR, Collins JH
JournalBiochemistry
PubMed ID10625482
In muscle thin filaments, the inhibitory region (residues 96-117) of troponin I (TnI) is thought to interact with troponin C (TnC) in the presence of Ca(2+) and with actin in the absence of Ca(2+). To better understand these interactions, we prepared mutant TnIs which contained a single Cys-96 or Cys-117 ... More
Proximity relationships between residue 6 of troponin I and residues in troponin C: further evidence for extended conformation of troponin C in the troponin complex.
AuthorsLuo Y, Leszyk J, Li B, Gergely J, Tao T
JournalBiochemistry
PubMed ID11112516
Skeletal muscle troponin C (TnC) adopts an extended conformation when crystallized alone and a compact one when crystallized with an N-terminal troponin I (TnI) peptide, TnI(1-47) [Vassylyev et al. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 4847-4852]. The N-terminal region of TnI (residues 1-40) was suggested to play a functional ... More
Peroxynitrite modification of protein thiols: oxidation, nitrosylation, and S-glutathiolation of functionally important cysteine residue(s) in the sarcoplasmic reticulum Ca-ATPase.
AuthorsViner RI, Williams TD, Schöneich C
JournalBiochemistry
PubMed ID10493809
Skeletal muscle contraction and relaxation is efficiently modulated through the reaction of reactive oxygen-nitrogen species with sarcoplasmic reticulum protein thiols in vivo. However, the exact locations of functionally important modifications are at present unknown. Here, we determine by HPLC-MS that the modification of one (out of 24) Cys residue of ... More
Internal movement in myosin subfragment 1 detected by fluorescence resonance energy transfer.
AuthorsXing J, Cheung HC
JournalBiochemistry
PubMed ID7756279
We have determined intersite distances from Cys374 of actin to Cys707 (SH1) and Cys697 (SH2) of myosin subfragment 1 (S1) in actosubfragment 1 (A.S1) by fluorescence resonance energy transfer for rigor complex A.S1 and complexes containing bound ADP and ADP plus orthovanadate (Vi), A.S1.ADP, and A.S1.ADP.Vi. A single energy acceptor ... More
Resonance energy transfer between points in a reconstituted skeletal muscle thin filament. A conformational change of the thin filament in response to a change in Ca2+ concentration.
AuthorsMiki M
JournalEur J Biochem
PubMed ID2105212
The spatial relationships between Lys-61, Cys-374 on actin or SH1 on myosin subfragment-1 (S1) and Cys-190 on tropomyosin or Cys-133 on troponin-I (TnI) in a reconstituted thin filament were studied by fluorescence resonance energy transfer. 5-(2-Iodoacetylaminoethyl)aminonaphthalene 1-sulfonic acid (IAEDANS) attached to Lys-190 on tropomyosin or to Cys-133 on TnI was ... More
Ca(2+)-dependent, myosin subfragment 1-induced proximity changes between actin and the inhibitory region of troponin I.
AuthorsKobayashi T, Kobayashi M, Collins JH
JournalBiochim Biophys Acta
PubMed ID11690651
In order to help understand the spatial rearrangements of thin filament proteins during the regulation of muscle contraction, we used fluorescence resonance energy transfer (FRET) to measure Ca(2+)-dependent, myosin-induced changes in distances and fluorescence energy transfer efficiencies between actin and the inhibitory region of troponin I (TnI). We labeled the ... More
Phosphorylation-dependent changes in the spatial relationship between Ca-ATPase polypeptide chains in sarcoplasmic reticulum membranes.
AuthorsBigelow DJ, Squier TC, Inesi G
JournalJ Biol Chem
PubMed ID1532393
In order to investigate possible structural changes associated with the coupling mechanisms of the Ca-ATPase in sarcoplasmic reticulum membranes, we have utilized fluorescence resonance energy transfer between spectroscopic probes covalently bound to different domains of the ATPase. Using time-correlated single photon counting, we have directly measured the energy transfer efficiency ... More
Chemical modification of the Ca(2+)-ATPase of rabbit skeletal muscle sarcoplasmic reticulum: identification of sites labeled with aryl isothiocyanates and thiol-directed conformational probes.
AuthorsWawrzynów A, Collins JH
JournalBiochim Biophys Acta
PubMed ID8218393
The Ca(2+)-ATPase protein of rabbit skeletal muscle sarcoplasmic reticulum is a single polypeptide chain of 1001 amino-acid residues. Among these residues are 24 Cys, 9 of which have previously been shown to be accessible to one or more thiol-specific reagents. Many studies on the structure and function of this Ca(2+)-ATPase ... More
Frequency-domain fluorescence spectroscopy resolves the location of maleimide-directed spectroscopic probes within the tertiary structure of the Ca-ATPase of sarcoplasmic reticulum.
AuthorsBigelow DJ, Inesi G
JournalBiochemistry
PubMed ID1825607
We have used fluorescence spectroscopy to characterize three covalently bound spectroscopic maleimide derivatives with respect to their location within the tertiary structure of the Ca-ATPase of sarcoplasmic reticulum (SR). These derivatives include (1) 2-(4'-maleimidoanilino)naphthalene-6-sulfonic acid, (2) 4-(dimethylamino)azobenzene-4'-maleimide, and (3) fluorescein 5'-maleimide. Biochemical assays demonstrate that modification with any of these ... More
Orientation of actin monomer in the F-actin filament: radial coordinate of glutamine-41 and effect of myosin subfragment 1 binding on the monomer orientation.
AuthorsKasprzak AA, Takashi R, Morales MF
JournalBiochemistry
PubMed ID3166995
We have employed the method of radial distance measurements in order to orient the actin monomer in the F-actin filament. This method utilizes fluorescence resonance energy transfer measurements of the distance between two equivalent chemical points located on two different monomers. The interprobe distance obtained this way is used to ... More
Detection of conformational changes in actin by fluorescence resonance energy transfer between tyrosine-69 and cysteine-374.
AuthorsMiki M
JournalBiochemistry
PubMed ID1932011
The distance between 5-(dimethylamino)naphthalene-1-sulfonyl chloride (dansyl chloride or DNS-Cl) attached to Tyr-69 and N-[[4-[4-(dimethylamino)phenyl]azo]phenyl]maleimide (DABMI) or N-[4-(dimethylamino)-3,5-dinitrophenyl]maleimide (DDPM) attached to Cys-374 in an actin monomer was measured to be 2.51 nm or 2.27 +/- 0.04 nm, respectively, by fluorescence resonance energy transfer. This distance does not change significantly when the ... More
Troponin-C-mediated calcium-sensitive changes in the conformation of troponin I detected by pyrene excimer fluorescence.
AuthorsStrasburg GM, Leavis PC, Gergely J
JournalJ Biol Chem
PubMed ID3965454
Troponin I (TnI) from rabbit white skeletal muscle was labeled at cysteines 48 and 64 with the fluorescent reagent N-(1-pyrene)maleimide. The fluorescence spectra of pyrene-labeled TnI (pyr-TnI) exhibit peaks characteristic of pyrene in its monomeric form and an additional peak resulting from formation of excited dimers (excimers), indicating that the ... More
Location of a contact site between actin and myosin in the three-dimensional structure of the acto-S1 complex.
AuthorsKasprzak AA, Chaussepied P, Morales MF
JournalBiochemistry
PubMed ID2532548
Using fluorescence resonance energy transfer (FRET), we measured distances from chromophores located at or near the actin-binding stretch of amino acids 633-642 of myosin subfragment 1 (S1), to five points in the acto-S1 complex. Specific labeling of this site was achieved by first attaching the desired chromophore to an "antipeptide" ... More
Dopamine biosynthesis is regulated by S-glutathionylation. Potential mechanism of tyrosine hydroxylast inhibition during oxidative stress.
AuthorsBorges CR, Geddes T, Watson JT, Kuhn DM
JournalJ Biol Chem
PubMed ID12376535
Tyrosine hydroxylase (TH), the initial and rate-limiting enzyme in the biosynthesis of the neurotransmitter dopamine, is inhibited by the sulfhydryl oxidant diamide in a concentration-dependent manner. The inhibitory effect of diamide on TH catalytic activity is enhanced significantly by GSH. Treatment of TH with diamide in the presence of [(35)S]GSH ... More
Models of the actin monomer and filament from fluorescence resonance-energy transfer.
AuthorsO'Donoghue SI, Hambly BD, dos Remedios CG
JournalEur J Biochem
PubMed ID1572360
We have developed algorithms for combining fluorescence resonance-energy transfer (FRET) efficiency measurements into structural models which predict the relative positions of the chemical groups used in FRET. We used these algorithms to construct models of the actin monomer and filament derived solely from FRET measurements based on seven distinct loci. ... More
Troponin T and Ca2+ dependence of the distance between Cys48 and Cys133 of troponin I in the ternary troponin complex and reconstituted thin filaments.
AuthorsLuo Y, Wu JL, Gergely J, Tao T
JournalBiochemistry
PubMed ID9283095
Contraction of vertebrate striated muscle is regulated by the interaction of Ca2+ with the heterotrimeric protein troponin (Tn), composed of troponin-C (TnC), troponin-I (TnI), and troponin-T (TnT). Although much is known about the Ca2+-induced conformational changes in TnC, the Ca2+-binding subunit of Tn, little is known about how TnI, the ... More
An extended conformation of calmodulin induces interactions between the structural domains of adenylyl cyclase from Bacillus anthracis to promote catalysis.
AuthorsDrum CL, Yan SZ, Sarac R, Mabuchi Y, Beckingham K, Bohm A, Grabarek Z, Tang WJ
JournalJ Biol Chem
PubMed ID10926933
The edema factor exotoxin produced by Bacillus anthracis is an adenylyl cyclase that is activated by calmodulin (CaM) at resting state calcium concentrations in infected cells. A C-terminal 60-kDa fragment corresponding to the catalytic domain of edema factor (EF3) was cloned, overexpressed in Escherichia coli, and purified. The N-terminal 43-kDa ... More
A new method for the selective isolation of cysteine-containing peptides. Specific labelling of the thiol group with a hydrophobic chromophore.
AuthorsChang JY, Knecht R, Braun DG
JournalBiochem J
PubMed ID6409087
A new method for the selective isolation of cysteine-containing peptides was designed. The method is based on the specific labelling of thiol groups with a hydrophobic chromophore followed by enzymic fragmentation of the labelled protein and reversed-phase high-pressure liquid-chromatographic separation of the peptide mixture. This new method has several distinct ... More
Energy-transfer measurements of the Cys35-Cys84 distance in bovine cardiac troponin C.
AuthorsLiou YM, Fuchs F
JournalBiochim Biophys Acta
PubMed ID8373830
Bovine cardiac troponin C (cTnC) has cysteine residues located in the non-functional Ca(2+)-binding loop I (Cys-35) and at the N-terminal end of the central helix (Cys-84) near site II, the regulatory Ca(2+)-binding site. Recently, we reported that the excimer fluorescence resulting from the dimerization of adjacent pyrene groups attached to ... More