Abstract:
:A detailed kinetic analysis of the catalytic trimer of aspartate transcarbamoylase containing the active site substitution H134A was performed to investigate the role of His 134 in the catalytic mechanism. Replacement of histidine by alanine resulted in decreases in the affinities for the two substrates, carbamoyl phosphate and aspartate, and the inhibitor succinate, by factors of 50, 10, and 6, respectively, and yielded a maximum velocity that was 5% that of the wild-type enzyme. However, the pK values determined from the pH dependence of the kinetic parameters, log V and log (V/K) for aspartate, the pK(i) for succinate, and the pK(ia) for carbamoyl phosphate, were similar for both the mutant and the wild-type enzymes, indicating that the protonated form of His 134 does not participate in binding and catalysis between pH 6.2 and 9.2. 13C and 15N isotope effects were studied to determine which steps in the catalytic mechanism were altered by the amino acid substitutions. The 13(V/K) for carbamoyl phosphate exhibited by the catalytic trimer containing alanine at position 134 revealed an isotope effect of 4.1%, probably equal to the intrinsic value and, together with quantitative analysis of the 15N isotope effects, showed that formation of the tetrahedral intermediate is rate-determining for the mutant enzyme. Thus, His 134 plays a role in the chemistry of the reaction in addition to substrate binding. The initial velocity pattern for the reaction catalyzed by the H134A mutant intersected to the left of the vertical axis, negating an equilibrium ordered kinetic mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)
journal_name
Biochemistryjournal_title
Biochemistryauthors
Waldrop GL,Turnbull JL,Parmentier LE,O'Leary MH,Cleland WW,Schachman HKdoi
10.1021/bi00143a031subject
Has Abstractpub_date
1992-07-21 00:00:00pages
6585-91issue
28eissn
0006-2960issn
1520-4995journal_volume
31pub_type
杂志文章相关文献
BIOCHEMISTRY文献大全abstract::Hepatic lipase (HL) hydrolysis of phosphatidylcholine (PC) was studied in recombinant high-density lipoprotein particles (r-HDL). r-HDL were made from cholate mixed micelles that contained PC, apo AI, and, in some cases, unesterified cholesterol. r-HDL were characterized using chemical composition, nondenaturing gradi...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi970356w
更新日期:1997-10-07 00:00:00
abstract::Three activities on the pathway of purine biosynthesis de novo in chicken liver, namely, glycinamide ribonucleotide synthetase, glycinamide ribonucleotide transformylase, and aminoimidazole ribonucleotide synthetase, have been found to reside on the same polypeptide chain. Three diverse purification schemes, utilizing...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00346a006
更新日期:1985-12-03 00:00:00
abstract::Vertical fluctuations of the terminal methyl groups of stearic acid acyl chains toward the surface of dimyristoylphosphatidylcholine (DMPC) bilayers have been investigated by using spin-label electron-electron double-resonance ( ELDOR ) methodology. Spin-label pairs consisting of two populations of stearic acid spin-l...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00305a032
更新日期:1984-05-08 00:00:00
abstract::[15N]Glycine was biosynthetically incorporated into normal cellular N-ras p21 and a position 12 transforming mutant, in order to produce p21 proteins containing several site-specific NMR probes at or near activating positions in the guanine nucleotide binding domain. We have previously assigned all five glycine resona...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00450a035
更新日期:1989-11-28 00:00:00
abstract::The crystal structure and spectroscopic properties of the periplasmic penta-heme cytochrome c nitrite reductase (NrfA) of Escherichia coli are presented. The structure is the first for a member of the NrfA subgroup that utilize a soluble penta-heme cytochrome, NrfB, as a redox partner. Comparison to the structures of ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi015765d
更新日期:2002-03-05 00:00:00
abstract::Endonuclease V of bacteriophage T4 binds to UV-irradiated deoxyribonucleic acid (DNA) but not to unirradiated DNA. We have developed an assay to detect this binding, based on the retention of enzyme--DNA complexes on nitrocellulose filters. The amount of complex retained, ascertained by using radioactive DNA, is a mea...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00549a026
更新日期:1980-04-15 00:00:00
abstract::Phospholipase B1 (PLB1), secreted by the pathogenic yeast Cryptococcus neoformans, has an established role in virulence. Although the mechanism of its phospholipase B, lysophospholipase, and lysophospholipase transacylase activities is unknown, it possesses lipase, subtilisin protease aspartate, and phospholipase moti...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi7009508
更新日期:2007-09-04 00:00:00
abstract::The RmFixL-RmFixJ oxygen signal transduction system ensures that a cascade of the Sinorhizobium meliloti nitrogen fixation genes is induced as the concentration of O2 drops below 50 microM in symbiotic nodules. Deoxy-RmFixL is a histidine protein kinase that catalyzes a phosphoryl transfer from ATP to the aspartate 54...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi051661h
更新日期:2005-11-22 00:00:00
abstract::Transmembrane chemoreceptors, also known as methyl-accepting chemotaxis proteins (MCPs), translate extracellular signals into intracellular responses in the bacterial chemotaxis system. MCP ligand binding domains control the activity of the CheA kinase, situated approximately 200 A away, across the cytoplasmic membran...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi801727m
更新日期:2009-03-10 00:00:00
abstract::The fluorescence polarization intensities from fluorescent probes and the electron paramagnetic resonance spectra from spin probes, specifically modifying elements of a biological assembly such as myosin sulfhydryl 1 (SH1) in muscle fibers, are interpreted in terms of probe order parameters using a model-independent m...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00116a029
更新日期:1992-01-14 00:00:00
abstract::Photosystem II catalyzes the light-driven oxidation of water and reduction of plastoquinone in oxygenic photosynthesis. The manganese stabilizing protein (MSP) of photosystem II is an extrinsic subunit that plays an important role in catalytic activity. This subunit can be extracted and re-bound to the photosystem II ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi9724467
更新日期:1998-04-21 00:00:00
abstract::An endo-beta-galactosidase, which hydrolyzes the internal beta-galactosidic linkages of R----GlcNAc (or GalNAc) beta 1----3Gal beta 1----4GlcNAc (or Glc), was isolated from the culture supernatant of Diplococcus pneumoniae. The enzyme, named endo-beta-galactosidase DII, hydrolyzed linear N-acetyllactosamine repeating ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00330a008
更新日期:1985-04-23 00:00:00
abstract::The clathrin assembly protein complex AP-2 is a multimeric subunit complex consisting of two 100-115-kDa subunits known as alpha and beta and 50- and 16-kDa subunits. The subunits have been dissociated and separated by ion-exchange chromatography in 7.5 M urea. Fractions highly enriched in either the alpha or beta sub...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00236a036
更新日期:1991-06-04 00:00:00
abstract::The protective effect of cyclic stable nitroxide free radicals, having SOD-like activity, against oxidative damage was studied by using Escherichia coli xthA DNA repair-deficient mutant hypersensitive to H2O2. Oxidative damage induced by H2O2 was assayed by monitoring cell survival. The metal chelator 1,10-phenanthrol...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00216a033
更新日期:1991-01-15 00:00:00
abstract::Site-specific mutagenesis has been used to create two mutant versions of aspartate transcarbamoylase. Arg-167 and Gln-231, both previously identified as interacting with the portion of the bisubstrate analogue N-(phosphonoacetyl)-L-aspartate (PALA) that corresponds to aspartate [Krause, K. L., Voltz, K. W., & Lipscomb...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00468a003
更新日期:1990-04-24 00:00:00
abstract::Purple membranes of Halobacterium halobium were studied by differential scanning calorimetry. No transition was detected at temperatures below 70 degrees C. A small endothermic transition was seen at about 80 degrees C and a larger one at 100 degrees C. The larger transition is the irreversible denaturation of bacteri...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00598a026
更新日期:1978-03-07 00:00:00
abstract::The structural protein nonerythroid alpha spectrin (alphaIISp) plays a role in the repair of DNA interstrand cross-links and is deficient in cells from patients with Fanconi anemia (FA), in which there is a defect in ability to repair such cross-links. We have proposed a model in which alphaIISp, whose stability is de...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi801483u
更新日期:2009-01-20 00:00:00
abstract::Deuterium magnetic resonance spectra (55.26 MHz) of cholesterol-3 alpha-d1 and epicholesterol-3 beta-d1 in dipalmitoylglycerophosphocholine (DPPC) liposomes were measured as a function of sterol-to-phospholipid ratio below (24 degrees C) and above (60 degrees C) the phase transition temperature of DPPC. From the quadr...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00353a017
更新日期:1986-03-11 00:00:00
abstract::Cryospectrokinetic studies of zinc and cobalt carboxypeptidase A disclosed two intermediates in the hydrolysis of both peptides and depsipeptides and furnished all the rate and equilibrium constants for the reaction scheme E + S in equilibrium ES1 in equilibrium ES2---E + P [Auld, D. S., Galdes, A., Geoghegan, K. F., ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00351a020
更新日期:1986-02-11 00:00:00
abstract::In this work, we show that adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S) is a substrate for yeast inorganic pyrophosphatase (PPase) (EC 3.6.1.1) and further, using chirally labeled [gamma-17O,18O]ATP gamma S, that enzyme-catalyzed hydrolysis to produce chiral inorganic thio[17O,18O]phosphate proceeds with inversio...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00300a002
更新日期:1984-02-28 00:00:00
abstract::Glutamate151 (E151) has been shown to be catalytically essential for the aminopeptidase from Vibrio proteolyticus (AAP). E151 acts as the general acid/base during the catalytic mechanism of peptide hydrolysis. However, a glutamate residue is not the only residue capable of functioning as a general acid/base during cat...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi0505823
更新日期:2005-09-13 00:00:00
abstract::While the iron-molybdenum cofactor (FeMoco) of nitrogenase, a constituent of the active site for nitrogen reduction, can be extracted into N-methylformamide (NMF) and pyrrollidinone, the inability to solubilize it in any other organic solvents has hampered further understanding of its structure and chemical properties...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00450a024
更新日期:1989-11-28 00:00:00
abstract::The structures of hemes a and a3 of maize and wheat germ cytochrome c oxidase were investigated by resonance Raman spectroscopy. Comparison between the plant and mammalian cytochrome oxidases revealed that (i) the vinyl groups associated with hemes a and a3 vibrate at higher frequencies in the plant enzyme than in the...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00489a028
更新日期:1990-09-18 00:00:00
abstract::By following the kinetics of abortive and productive synthesis in single-round transcription assays, we confirm the existence of two general classes of initial transcribing complexes (ITCs), which we term "productive ITC" and "unproductive ITC". The productive ITCs are able to escape from the promoter rapidly to produ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi0269613
更新日期:2003-04-08 00:00:00
abstract::Clustering of anionic lipids away from zwitterionic ones by cationic antimicrobial agents has recently been established as a mechanism of action of natural small, flexible peptides as well as non-natural synthetic peptide mimics. One of the largest classes of antimicrobial peptides consists of peptides that form catio...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi100378m
更新日期:2010-05-18 00:00:00
abstract::Myc and Max belong to the b-HLH-LZ family of transcription factors. Heterodimerization between Myc and Max or homodimerization of Max allows these proteins to bind their cognate DNA sequence known as the E-box (CACGTG). Recent evidence has suggested that the c-Myc/Max heterodimeric b-HLH-LZ could interact to form a he...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi700076m
更新日期:2007-09-11 00:00:00
abstract::Our recent demonstration that DNA polymerase X (Pol X), the DNA repair polymerase encoded by the African swine fever virus (ASFV), is extremely error prone during single-nucleotide gap filling led us to hypothesize that it might contribute to genetic variability in ASFV. For the infidelity of Pol X to be relevant, how...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi047706g
更新日期:2005-06-14 00:00:00
abstract::In order to quantitate the contributions of the polar, uncharged amino acids to the stability of the native state of staphylococcal nuclease, each of the 13 alanines, 9 glycines, 9 threonines, 6 prolines, 6 glutamines, 6 asparagines, and 3 serines was substituted, either with both alanine and glycine or with 1 of thes...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00140a005
更新日期:1992-06-30 00:00:00
abstract::The binding of the codon UUC to the isolated anticodon loop of tRNAPhe (yeast) has been studied as a model of codon recognition by a simple adaptor. Fluorescence titrations demonstrate that UUC binds to the isolated anticodon loop with an equilibrium constant of 1.4 X 10(3) M-1 (at 7.2 degrees C). Equilibrium sediment...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00369a005
更新日期:1986-10-21 00:00:00
abstract::This paper reports on a solution NMR characterization of the sequence selectivity and metal ion specificity in chromomycin-DNA oligomer complexes in the presence of divalent cations. The sequence selectivity studies have focused on chromomycin complexes with the self-complementary d(A1-A2-G3-G4-C5-C6-T7-T8) duplex con...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00501a012
更新日期:1990-12-11 00:00:00