Co-expression of chemotactic ligand receptors on human peripheral blood monocytes.
AuthorsOhura K, Katona IM, Wahl LM, Chenoweth DE, Wahl SM
JournalJ Immunol
PubMed ID2435802
'Directed migration of monocytes is dependent upon interaction of cell surface receptors and specific chemotactic ligands. To determine whether circulating human monocytes express multiple chemotactic ligand receptors or whether subpopulations of monocytes exist with a single receptor specificity, nonoverlapping fluorescent probes for two chemotactic ligands, N-formyl methionyl leucyl phenylalanine (FMLP) ... More
Glycophorin A helical transmembrane domains dimerize in phospholipid bilayers: a resonance energy transfer study.
AuthorsAdair BD, Engelman DM
JournalBiochemistry
PubMed ID8180176
'Glycophorin A and its isolated transmembrane region (GpATM) are each known to form sequence-specific dimers in SDS micelles. Whether this behavior accurately reflects behavior in red cell membranes or lipid bilayers, however, has remained unclear. Resonance energy transfer between labeled GpATM peptides has been used to observe dimerization of GpATM ... More
Active site-independent recognition of substrates and product by bovine prothrombinase: a fluorescence resonance energy transfer study.
AuthorsBoskovic DS, Troxler T, Krishnaswamy S
JournalJ Biol Chem
PubMed ID14988397
'The conversion of prothrombin to thrombin is catalyzed by prothrombinase, an enzyme complex composed of the serine proteinase factor Xa and a cofactor protein, factor Va, assembled on membranes. Kinetic studies indicate that interactions with extended macromolecular recognition sites (exosites) rather than the active site of prothrombinase are the principal ... More
Factor IXa protects factor VIIIa from activated protein C. Factor IXa inhibits activated protein C-catalyzed cleavage of factor VIIIa at Arg562.
'Factor VIIIa is inactivated by both factor IXa and activated protein C. The latter protease rapidly attacked a site at Arg562 (A2 subunit), whereas both proteases slowly cleaved factor VIIIa at Arg336 (A1 subunit). Cofactor inactivation catalyzed by activated protein C was 8-fold faster than that catalyzed by factor IXa. ... More
Surface density determination in membranes by fluorescence energy transfer.
AuthorsFung BK, Stryer L
JournalBiochemistry
PubMed ID728398
Position-specific incorporation of dansylated non-natural amino acids into streptavidin by using a four-base codon.
AuthorsHohsaka T, Muranaka N, Komiyama C, Matsui K, Takaura S, Abe R, Murakami H, Sisido M
JournalFEBS Lett
PubMed ID14988018
Novel non-natural amino acids carrying a dansyl fluorescent group were designed, synthesized, and incorporated into various positions of streptavidin by using a CGGG four-base codon in an Escherichia coli in vitro translation system. 2,6-Dansyl-aminophenylalanine (2,6-dnsAF) was found to be incorporated into the protein more efficiently than 1,5-dansyl-lysine, 2,6-dansyl-lysine, and 1,5-dansyl-aminophenylalanine. ... More
Fluorescent hydrazides for the high-performance liquid chromatographic determination of biological carbonyls.
AuthorsAnderson JM
JournalAnal Biochem
PubMed ID2937341
Methods for the determination of carbonyl compounds of biological origin by high-performance liquid chromatography were improved by the use of new fluorescent derivatizing agents. Eight fluorescent hydrazides were either synthesized or obtained commercially and compared to dansyl hydrazine (1-dimethylaminonaphthalene-5-sulfonylohydrazide). Four of the compounds yielded carbonyl hydrazones with a higher relative ... More
Time-Resolved Fluorescence Energy Transfer DNA Helicase Assays for High Throughput Screening.
AuthorsEarnshaw DL, Moore KJ, Greenwood CJ, Djaballah H, Jurewicz AJ, Murray KJ, Pope AJ
JournalJ Biomol Screen
PubMed ID10838444
DNA helicases are responsible for the unwinding of double-stranded DNA, facilitated by the binding and hydrolysis of 5'-nucleoside triphosphates. These enzymes represent an important class of targets for the development of novel anti-infective agents particularly because opportunity exists for synergy with existing therapies targeted at other enzymes involved in DNA ... More
A fluorescence energy transfer method for analyzing protein oligomeric structure: application to phospholamban.
AuthorsLi M, Reddy LG, Bennett R, Silva ND, Jones LR, Thomas DD
JournalBiophys J
PubMed ID10233073
We have developed a method using fluorescence energy transfer (FET) to analyze protein oligomeric structure. Two populations of a protein are labeled with fluorescent donor and acceptor, respectively, then mixed at a defined donor/acceptor ratio. A theoretical simulation, assuming random mixing and association among protein subunits in a ring-shaped homo-oligomer, ... More
Zymogen/enzyme discrimination using peptide chloromethyl ketones.
AuthorsWilliams EB, Krishnaswamy S, Mann KG
JournalJ Biol Chem
PubMed ID2708377
Glutamylglycinylarginyl chloromethyl ketone, tyrosylglycinylarginyl chloromethyl ketone, and phenylalanylprolylarginyl chloromethyl ketone have been labeled at their amino termini using fluorescein, rhodamine-X, lissamine-rhodamine, pyrene, and the 1,5-, 2,5-, and 2,6-dimethylaminonaphthalene-1-sulfonyl moieties. These peptidyl chloromethyl ketones have also been modified by incorporation of biotin and epsilon-amino caproyl biotin. The ability of these various ... More
The binding of activated protein C to factors V and Va.
AuthorsKrishnaswamy S, Williams EB, Mann KG
JournalJ Biol Chem
PubMed ID3755431
Activated protein C has been derivatized with the active site-directed fluorophore 2-(dimethylamino)-6-naphthalenesulfonylglutamylglycylarginyl chloromethyl ketone (2,6-DEGR-APC). Covalently modified activated protein C has been used to investigate the binding interactions of the protein to factors V and Va in the presence of phospholipid vesicles. The fluorescence polarization of the 6-dimethylaminonaphthalene-2-sulfonyl moiety increased ... More