Abstract:
:The prothrombinase complex assembles through reversible interactions between factor Xa, factor Va and acidic phospholipid-containing membranes in the presence of calcium ions. This complex catalyses the conversion of prothrombin to thrombin through two proteolytic steps. We have used prethrombin 2 as a substrate analog for the first cleavage reaction of prothrombin activation (cleavage at Arg323-Ile324) catalyzed by the prothrombinase complex and have also relied on the known ability of prethrombin 2 to interact tightly but reversibly with fragment 2 or fragment 1.2. The kinetics of cleavage at Arg323-Ile324 have been assessed with these substrate analogs to investigate the contribution of cofactor-substrate interactions mediated by the fragment 2 domain to the ability of factor Va to enhance the catalytic efficiency of factor Xa within the prothrombinase complex. Initial velocity measurements indicated that the rate of prethrombin 2 cleavage by the factor Xa-PCPS binary complex was increased by a factor of approximately 1300 upon the addition of saturating concentrations of factor Va to assemble prothrombinase. Although the measured initial velocity was higher when either fragment 2 or fragment 1.2 was present, the factor Va-dependent enhancement in initial rate (2600- and 1500-fold) was comparable in each case. Steady state kinetic constants were obtained using prethrombin 2, prethrombin 2 plus fragment 2, and prethrombin 2 plus fragment 1.2 as substrates. For each substrate, the addition of saturating concentrations of factor Va to the Xa-PCPS binary complex led to increases in catalytic efficiency of between 1000 and 9000-fold. The kcat/Km for prethrombin 2 cleavage by prothrombinase was essentially identical to that obtained for prethrombin 2 saturated with fragment 2. Thus, comparable accelerating effects of factor Va are observed independent of the presence of the fragment 2 domain in the substrate. The results indicate that interactions between factor Va and the substrate mediated by the fragment 2 domain do not contribute in a dominant way to the ability of factor Va to enhance the catalytic efficiency of factor Xa within the prothrombinase complex.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Krishnaswamy S,Walker RKdoi
10.1021/bi9623993subject
Has Abstractpub_date
1997-03-18 00:00:00pages
3319-30issue
11eissn
0006-2960issn
1520-4995pii
bi9623993journal_volume
36pub_type
杂志文章相关文献
BIOCHEMISTRY文献大全abstract::The glucose derivative, 2,2,6,6-tetramethylpiperidine-1-oxylglucose (TEMPO-glucose) was synthesized and examined for its ability to substitute for glucose as a substrate for the intestinal brush border membrane Na+/glucose cotransporter. TEMPO-glucose inhibited Na(+)-dependent phlorizin binding with an apparent KI of ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00231a012
更新日期:1991-04-30 00:00:00
abstract::New isoforms of CYP51 (sterol 14alpha-demethylase), an essential enzyme in sterol biosynthesis and primary target of azole antimycotic drugs, are found in pathogenic protists, Trypanosoma brucei(TB), T. vivax, T. cruzi, and Leishmania major. The sequences share approximately 80% amino acid identity and are approximate...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi048967t
更新日期:2004-08-24 00:00:00
abstract::Protein amyloid fibrils are originally identified as pathological entities in a variety of neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease. Recent studies have revealed that amyloid fibrils also serve as functional protein assemblies to fulfill a wide range of biological functions. Decip...
journal_title:Biochemistry
pub_type: 杂志文章,评审
doi:10.1021/acs.biochem.9b01069
更新日期:2020-02-11 00:00:00
abstract::We have used resonance energy transfer to read out the interactions of the alpha subunit of transducin (alpha T) with the transducin beta gamma subunit complex (beta gamma T) and to compare the rate of aluminum fluoride-induced alpha T activation, as reflected by the enhancement of the alpha T tryptophan fluorescence,...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00199a046
更新日期:1994-08-23 00:00:00
abstract::Flavocytochrome b2 from Saccharomyces cerevisiae couples L-lactate dehydrogenation to cytochrome c reduction. At 25 degrees C, 0.10 M ionic strength, and saturating L-lactate concentration, the turnover rate is 207 s-1 [per cytochrome c reduced; Miles, C. S., Rouviere, N., Lederer, F., Mathews, F. S., Reid, G. A., Bla...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi9522561
更新日期:1996-05-21 00:00:00
abstract::Mycobacterium tuberculosis and many other members of the Actinomycetes family produce mycothiol, i.e., 1-d-myo-inosityl-2-(N-acetyl-l-cysteinyl)amido-2-deoxy-alpha-d-glucopyranoside (MSH or AcCys-GlcN-Ins), to act against oxidative and antibiotic stress. The biosynthesis of MSH is essential for cell growth and has bee...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi7011492
更新日期:2007-10-09 00:00:00
abstract::The crystal structure of the catalytic domain of rat DNA polymerase beta revealed that Asp256 is located in proximity to the previously identified active site residues Asp190 and Asp192. We have prepared and kinetically characterized the nucleotidyl transfer activity of wild type and several mutant forms of human and ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00049a008
更新日期:1995-12-12 00:00:00
abstract::A receptor binding region of mouse interferon gamma (IFN gamma) has previously been localized to the N-terminal 39 amino acids of the molecule by use of synthetic peptides and monoclonal antibodies. In this report, a detailed analysis of the synthetic peptide corresponding to this region, IFN gamma (1-39), is presente...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00237a022
更新日期:1991-06-11 00:00:00
abstract::The alpha-helix formed by the amino acid residues 9-19 of the B-chain of insulin is involved in the stabilization of its three-dimensional structure. We have shown that modification at positions B9, B10, B12, and B16 results in analogues possessing biological activities ranging from ca. 0.2% to ca. 500% relative to th...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00061a022
更新日期:1993-03-16 00:00:00
abstract::We have investigated the binding of a pure H1 histone, the mouse variant H1e, to a 214 bp fragment of DNA from pBR322. Binding was monitored by observing the effects of the protein on melting of the DNA and by a gel-mobility-shift assay. We found, using this highly purified system, that H1e protein binds preferentiall...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00167a049
更新日期:1994-01-11 00:00:00
abstract::Corticosteroid binding globulin (CBG), a serum glycoprotein which binds glucocorticoids and progestins with high affinity, is widely distributed throughout the animal world. Although its charge and size characteristics have largely been conserved across species, we found the behavior of CBG in squirrel monkey (Saimiri...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00407a046
更新日期:1988-04-05 00:00:00
abstract::DBA/2 mouse spleen cells were stimulated in vitro by (a) alloantigen (mitomycin-treated CBA/J splenocytes), (b) the T cell mitogen concanavalin A (Con A), and (c) the B cell mitogen lipopolysaccharide (LPS). The cultures were pulsed for 10 h with (14)C-labeled galactose and glucosamine. Radiolabeled glycosphingolipids...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00519a002
更新日期:1981-08-04 00:00:00
abstract::The reversible decline of the nicotinic acetylcholine receptor's response to acetylcholine during prolonged exposure to acetylcholine is known as desensitization. Here, we studied ethanol's modulation of fast agonist-induced desensitization of the nicotinic acetylcholine receptor in postsynaptic membrane vesicles from...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00197a009
更新日期:1994-08-09 00:00:00
abstract::Overall activation volumes for multistep reactions are not usually pressure independent. The present investigation gives a quantitative description of this effect under Theory. Simple relations are obtained which can easily be applied to experimental data and which allow more insight into the dynamics of enzyme reacti...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00405a024
更新日期:1988-03-08 00:00:00
abstract::Many early investigations on triple helices have been devoted to the study of the triplex formed by dT*dA-dT base triplets in which the third strand is oriented parallel to the dA strand. We now describe an intramolecular triple helix with dT*dA-dT base triplets in which the pyrimidine third strand is oriented antipar...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00045a023
更新日期:1995-11-14 00:00:00
abstract::Factor V and factor Va binding to single bilayer phospholipid vesicles was investigated by light-scattering intensity measurements. This technique allows the measurement of free and phospholipid-bound protein concentrations from which equilibrium constants can be obtained. As controls, the Ca2+-dependent phospholipid ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00587a023
更新日期:1979-10-02 00:00:00
abstract::Purine nucleoside phosphorylase (PNP) catalyzes the reversible phosphorolysis of purine ribonucleosides and 2'-deoxyribonucleosides, yielding the purine base and (2'-deoxy)ribose 1-phosphate as products. While this enzyme has been extensively studied, several questions with respect to the catalytic mechanism have rema...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/acs.biochem.5b01347
更新日期:2016-04-12 00:00:00
abstract::Proton NMR spectroscopy was applied to myoglobin in the ferric, water-liganded form (metMbH2O) and the apo form (apoMb) to probe the structure and stability of the latter. Proteins from sperm whale and horse skeletal muscles were studied to simplify the spectral assignment task. Nuclear Overhauser effects and the resp...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00143a018
更新日期:1992-07-21 00:00:00
abstract::The 1,2-diacyl-sn-glycerols (1,2-DGs) are the predominant naturally occurring isomer found in cell membranes, lipid droplets, and lipoproteins. They are involved in the metabolism of monoacylglycerols, triacylglycerols, and phospholipids. The 1,2-DGs participate in the activation of protein kinase C, in phosphorylatio...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00500a008
更新日期:1990-12-04 00:00:00
abstract::The topography of the photosynthetic reaction center (RC) polypeptides (H, M, and L) was investigated by proteolysis and radioiodination of membrane vesicles isolated from Rhodopseudomonas sphaeroides. Chromatophores, obtained from French-pressed cell lysates, are closed vesicles' and oriented inside out with respect ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00519a012
更新日期:1981-08-04 00:00:00
abstract::The Na+-translocating NADH:ubiquinone oxidoreductase (Na+-NQR) from Vibrio harveyi was purified and studied by EPR and visible spectroscopy. Two EPR signals in the NADH-reduced enzyme were detected: one, a radical signal, and the other a line around g = 1.94, which is typical for a [2Fe-2S] cluster. An E(m) of -267 mV...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi002545b
更新日期:2001-06-19 00:00:00
abstract::Nuclease footprinting techniques have been employed to investigate the interaction between the diuretic drug amiloride, a sodium channel blocker with potential therapeutic use in the treatment of cystic fibrosis, and three DNA fragments of defined sequence. Using either DNAse I or micrococcal nuclease as probes, an un...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00061a008
更新日期:1993-03-16 00:00:00
abstract::Copper binding to the human copper chaperone for superoxide dismutase (hCCS) has been investigated by X-ray absorption spectroscopy. Stoichiometry measurements on the dialyzed, as-isolated protein indicated that up to 3.5 Cu ions bound per hCCS molecule. Reduction with either sodium dithionite or dithiothreitol decrea...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi000690j
更新日期:2000-06-27 00:00:00
abstract::The immunoglobulin E-binding protein, epsilon BP (also known as CBP35, Mac-2, L-34, and L-29), is a beta-galactoside-binding protein of approximately 30 kDa and a member of the animal lectin family termed S-type or S-Lac. Multiple biological activities have been attributed to this lectin such as mediation of IgE bindi...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00186a038
更新日期:1994-05-24 00:00:00
abstract::Thiazolinyl imine reductases catalyze the NADPH-dependent reduction of a thiazoline to a thiazolidine, a required step in the formation of the siderophores yersiniabactin (Yersinia spp.) and pyochelin (Pseudomonas aeruginosa). These stand-alone nonribosomal peptide tailoring domains are structural homologues of sugar ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/acs.biochem.6b00735
更新日期:2016-09-27 00:00:00
abstract::The generation of multiprotein complexes at receptors and adapter proteins is crucial for the activation of intracellular signaling pathways. In this study, we used multiple biochemical and biophysical methods to examine the binding properties of several SH2 and SH3 domain-containing signaling proteins as they interac...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi0357311
更新日期:2004-04-13 00:00:00
abstract::Specificity of the binding of sterols and related compounds with purified F-protein (fusion protein) of the HVJ (Sendai virus) was studied by binding competition with [3H]cholesterol. Requirement for cholesterol or the A/B ring trans structure and nonrequirement for the 3-hydroxyl group were found in this binding. Bin...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00404a035
更新日期:1988-02-23 00:00:00
abstract::Secretory carrier membrane protein 2 (SCAMP2) functions in late steps of membrane fusion in calcium-dependent granule exocytosis. A basic/hydrophobic peptide segment within SCAMP2 (SCAMP2 E: CWYRPIYKAFR) has been implicated in this function and shown to bind and sequester phosphatidylinositol 4,5-bisphosphate [PI(4,5)...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi701121j
更新日期:2007-09-25 00:00:00
abstract::Biotin synthase, the enzyme which catalyzes the last step of the biosynthesis of biotin, contains only (2Fe-2S)(2+) clusters when isolated under aerobic conditions. Previous results showed that reduction by dithionite or photoreduced deazaflavin converts the (2Fe-2S)(2+) to (4Fe-4S)(2+,+). However, until now, no detai...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi034426c
更新日期:2003-07-29 00:00:00
abstract::The interaction of solvent of the substrate binding domain of the bacterial heat shock 70 chaperone protein DnaK was studied in its apo form and with bound hydrophobic substrate peptide, using refined nuclear magnetic resonance experiments. Distinct differences between the two states of the protein were observed. Acco...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi030097c
更新日期:2003-09-30 00:00:00