Abstract:
:Primary kinetic isotope effects on Vmax were observed in both reaction directions upon racemizing samples of [2-2H]glutamate with the cofactor-independent glutamate racemase from Lactobacillus. This supports a deprotonation/protonation mechanism for racemization in which the breaking of the carbon-hydrogen bond at C-2 is partially rate-determining. Substantial "overshoots" were observed when the time course of racemization of either enantiomer of glutamate was monitored using circular dichroism spectroscopy. This is consistent with a "two-base" mechanism accompanied by a kinetic isotope effect. "Competitive deuterium washout" experiments were used to measure kinetic isotope effects on Vmax/Km of 2.5 for (S)-glutamate and 3.4 for (R)-glutamate. The ratio of the notably different isotope effects was confirmed by "double competitive deuterium washout" experiments. Site-directed mutagenesis was used to generate the mutant C73A and C184A enzymes. In each case the mutant enzymes were inactive as racemases. The two mutant enzymes are, however, capable of catalyzing the elimination of HCl from opposite enantiomers of threo-3-chloroglutamic acid, a process that presumably requires only one enzymic base. This finding indicates that the active sites of the mutant enzymes are intact and that the two cysteines flank the bound substrate molecule. It appears that cysteine-73 is responsible for the abstraction of the C-2 hydrogen from (R)-glutamate and cysteine-184 abstracts the proton from (S)-glutamate in the racemization reaction of the wild-type enzyme.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Tanner ME,Gallo KA,Knowles JRdoi
10.1021/bi00066a021subject
Has Abstractpub_date
1993-04-20 00:00:00pages
3998-4006issue
15eissn
0006-2960issn
1520-4995journal_volume
32pub_type
杂志文章相关文献
BIOCHEMISTRY文献大全abstract::DNA transaction reactions require formation of nucleoprotein complexes that involve multifaceted DNA-protein and protein-protein interactions. Genetic and biochemical studies suggested that the higher order Gal repressosome structure, which governs the transcription of two tandem galpromoters in Escherichia coli, invo...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi047720t
更新日期:2005-04-12 00:00:00
abstract::The effect of hydrostatic pressure upon solutions of chymotrypsinogen and lysozyme at room temperature has been followed by employing a new technique [Chryssomallis, G. S., Drickamer, H. G., & Weber, G. (1978) J. Appl. Phys. 49, 3084] that permits the measurement of fluorescence polarization at pressures of up to 10 k...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00517a002
更新日期:1981-07-07 00:00:00
abstract::The unique characteristics of DNA charge transport (CT) have prompted an examination of roles for this chemistry within a biological context. Not only can DNA CT facilitate long-range oxidative damage of DNA, but redox-active proteins can couple to the DNA base stack and participate in long-range redox reactions using...
journal_title:Biochemistry
pub_type: 杂志文章,评审
doi:10.1021/bi501520w
更新日期:2015-02-03 00:00:00
abstract::The association of nucleolar phosphoprotein C23 with preribosomal ribonucleoprotein (RNP) particles was examined in Novikoff hepatoma nucleoli. RNA was labeled with [3H]uridine for various times in cell suspensions, and RNP particles were extracted from isolated nucleoli and fractionated by sucrose gradient ultracentr...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00368a063
更新日期:1986-10-07 00:00:00
abstract::The early steps of the proposed mechanistic pathway for dehydroquinate synthase have been probed with a series of substrate analogues. These analogues, 3-9, are structurally prohibited from undergoing the beta-elimination of inorganic phosphate that represents the committed step in the conversion of the substrate 3-de...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00445a010
更新日期:1989-09-19 00:00:00
abstract::The catalytic activity of protein-tyrosine phosphatases (PTPs) is mediated by a cysteine side chain which carries out a nucleophilic attack initiating the phosphate cleavage. Experimentally, it has been observed that the active site cysteine has a remarkably low pKa. In the present study, we have investigated the orig...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi971187i
更新日期:1998-04-21 00:00:00
abstract::RNA pur*pur-pyr and pyr*pur-pyr (pur = purine, pyr = pyrimidine) triple helices consisting of a Watson-Crick base-paired 28mer hairpin duplex and a Hoogsteen base-paired purine or pyrimidine 12mer are targeted with photoactivated cleavage by the metal complex Rh(phen)2phi3+ (phen = phenanthroline, phi = 9, 10-phenanth...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi980509v
更新日期:1998-06-23 00:00:00
abstract::Thermal transitions of type I collagen fibrils were investigated by differential scanning calorimetry and spectrophotometry of turbidity within a wide range of external conditions. The advanced microcalorimeter allowed us to carry out the measurements at low concentrations of collagen (0.15-0.3 mg/mL). At these concen...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi980360n
更新日期:1998-06-02 00:00:00
abstract::Post-translational phosphorylation and the related conformational changes in signaling proteins are responsible for regulating a wide range of subcellular processes. Human Pin1 is central to many of these cell signaling pathways in normal and aberrant subcellular processes, catalyzing cis-trans isomerization of the pe...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi2009954
更新日期:2011-11-08 00:00:00
abstract::Phosphotriesterase, as isolated from Pseudomonas diminuta, is capable of detoxifying widely used pesticides such as paraoxon and parathion and various mammalian acetylcholinesterase inhibitors. The enzyme requires a binuclear metal center for activity. Recently, the three-dimensional structure of the apoenzyme was sol...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00025a002
更新日期:1995-06-27 00:00:00
abstract::Phosphorus NMR spectroscopy is an important technique for the investigation of metabolism in tissues and intact organisms (including man). However, quantitation of the signals from an NMR experiment is difficult because it is not known from which regions of a cell metabolites are detected. It is generally believed tha...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00439a003
更新日期:1989-06-27 00:00:00
abstract::Biotin synthase, a member of the "radical SAM" family, catalyzes the final step of the biotin biosynthetic pathway, namely, the insertion of a sulfur atom into dethiobiotin (DTB). The active form of the enzyme contains two iron-sulfur clusters, a [4Fe-4S](2+) cluster liganded by Cys-53, Cys-57, and Cys-60 and the S-ad...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi060662m
更新日期:2006-10-10 00:00:00
abstract::Despite many studies suggesting the role of enolpyruvate as a bound intermediate in the pyruvate kinase reaction, direct evidence for it has been lacking. By use of a combination of chemical trapping and isolation of a derivative, significant amounts of enzyme-bound enolpyruvate have now been demonstrated. The method ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00217a022
更新日期:1991-01-22 00:00:00
abstract::The head of the P22 bacteriophage is interrupted by a unique dodecameric portal vertex that serves as a conduit for the entrance and exit of the DNA. Here, the in vitro unfolding/refolding processes of the portal protein of P22 were investigated at different temperatures (1, 25, and 37 degrees C) through the use of ur...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi700006d
更新日期:2007-06-26 00:00:00
abstract::Non-heme diiron clusters occur in a number of enzymes (e.g., ribonucleotide reductase, methane monooxygenase, and Delta9-stearoyl-ACP desaturase) that activate O2 for chemically difficult oxidation reactions. In each case, a kinetically labile peroxo intermediate is believed to form when O2 reacts with the diferrous e...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi981838q
更新日期:1998-10-20 00:00:00
abstract::We have employed high-performance liquid chromatography (HPLC) to investigate the syntheses of histones H1 and H1o as synchronized cells traverse from mitosis to S phase. Chinese hamster (line CHO) cells were synchronized by mitotic selection, and, at appropriate times, they were pulse labeled for 1 h with [3H]lysine....
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00329a031
更新日期:1985-04-09 00:00:00
abstract::The nonbulky 5',8-cyclopurine DNA lesions (cP) and the bulky, benzo[ a]pyrene diol epoxide-derived stereoisomeric cis- and trans- N2-guanine adducts (BPDE-dG) are good substrates of the human nucleotide excision repair (NER) mechanism. These DNA lesions were embedded at the In or Out rotational settings near the dyad ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/acs.biochem.8b01066
更新日期:2019-02-12 00:00:00
abstract::Thrombospondin-1 (TSP1) binding to calreticulin (CRT) on the cell surface signals focal adhesion disassembly, leading to the intermediate adhesive phenotype, cell migration, anoikis resistance, and collagen stimulation. Residues Lys 24 and 32 in TSP1 and amino acids 24-26 and 32-34 in CRT have been shown through bioch...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi101639y
更新日期:2011-02-01 00:00:00
abstract::The association between the papillomavirus E1 and E2 proteins is an important regulatory interaction, imparting coordinated control of viral transcription and replication. Using fluorescence polarization, we have characterized the interactions between HPV-11 E1, HPV-11 E2, and DNA in solution at equilibrium. For these...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi982616v
更新日期:1999-04-06 00:00:00
abstract::Based on the atomic structures of the mitochondrial cytochrome bc(1) complex, it has been proposed that the soluble domain of the [2Fe-2S] Rieske iron-sulfur protein (ISP) must rotate by ca. 60 degrees and translate through an appreciable distance between two binding sites, proximal to cytochrome c(1) and to the lumen...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi9921992
更新日期:2000-03-14 00:00:00
abstract::Comparative sequence analysis has successfully predicted secondary structure and tertiary interactions in ribosomal and other RNAs. Experiments presented here ask whether the scope of comparative sequence-based predictions can be extended to specific interactions between proteins and RNA, using as a system the well-ch...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi9826411
更新日期:1999-03-23 00:00:00
abstract::The third intracellular loop (IL3) of G protein-coupled receptors (GPCRs) is an important contact domain between GPCRs and their G proteins. Previously, the IL3 of Ste2p, a Saccharomyces cerevisiae GPCR, was suggested to undergo a conformational change upon activation as detected by differential protease susceptibilit...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi200254h
更新日期:2011-08-16 00:00:00
abstract::The VWF A1 domain seems to possess two heparin binding regions (residues 565-587 and 633-648) displaying positively charged amino acids, but the overall polyanion-A1 domain interaction scheme remains essentially elusive. To probe this molecular reaction as well as the role of electrostatic forces in VWF-heparin intera...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi020044f
更新日期:2002-05-28 00:00:00
abstract::Human insulin-like growth factors I and II (hIGF-I, hIGF-II) are potent stimulators of cell and growth processes. They display high sequence similarity to both the A and B chains of insulin but contain an additional connecting C-domain, which reflects their secretion without specific packaging or precursor conversion....
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi020084j
更新日期:2002-07-30 00:00:00
abstract::Transferrin exhibits heterogeneity in glycosylation characteristic of pathological changes in alcohol abuse and congenital disorders in glycosylation. This study investigated an alternative approach in the detection of carbohydrate-deficient transferrin based on the premise that glycosylation may afford some degree of...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi051749v
更新日期:2005-12-13 00:00:00
abstract::The bovine protein tyrosine phosphatase (BPTP) is a member of the class of low-molecular weight protein tyrosine phosphatases (PTPases) found to be ubiquitous in mammalian cells. The catalytic site of BPTP contains a CX(5)R(S/T) phosphate-binding motif or P-loop (residues 12-19) which is the signature sequence for all...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi990381x
更新日期:1999-09-07 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::Living organisms have evolved a vast array of catalytic functions that make them ideally suited for the production of medicinally and industrially relevant small-molecule targets. Indeed, native metabolic pathways in microbial hosts have long been exploited and optimized for the scalable production of both fine and co...
journal_title:Biochemistry
pub_type: 杂志文章,评审
doi:10.1021/bi200416g
更新日期:2011-06-21 00:00:00
abstract::The selenoamino acids methylselenocysteine (MeSeCys) and selenomethionine (SeMet) have disparate efficacies as anticancer agents. Herein, we use X-ray absorption spectroscopy to determine the chemical form of selenium in human neuroblastoma cells. Cells treated with MeSeCys contain a significant diselenide component, ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi201462u
更新日期:2012-01-24 00:00:00
abstract::An endogenous calcium-activated sulfhydryl protease in chick oviduct cytosol has been utilized to study the structure of the chick oviduct progesterone receptor subunits, progestophilins A (79 000 g/mol) and B (117 000 g/mol). The protease is not a normal component of the native progesterone receptor aggregate (6 and ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00543a015
更新日期:1980-01-22 00:00:00