Abstract:
:Binding of substrate to ornithine 4,5-aminomutase (OAM) and methylmalonyl-CoA mutase (MCM) leads to the formation of an electrostatic interaction between a conserved glutamate side chain and the adenosyl ribose of the adenosylcobalamin (AdoCbl) cofactor. The contribution of this residue (Glu338 in OAM from Clostridium sticklandii and Glu392 in human MCM) to AdoCbl Co-C bond labilization and catalysis was evaluated by substituting the residue with a glutamine, aspartate, or alanine. The OAM variants, E338Q, E338D, and E338A, showed 90-, 380-, and 670-fold reductions in catalytic turnover and 20-, 60-, and 220-fold reductions in k(cat)/K(m), respectively. Likewise, the MCM variants, E392Q, E392D, and E392A, showed 16-, 330-, and 12-fold reductions in k(cat), respectively. Binding of substrate to OAM is unaffected by the single-amino acid mutation as stopped-flow absorbance spectroscopy showed that the rates of external aldimine formation in the OAM variants were similar to that of the native enzyme. The decrease in the level of catalysis is instead linked to impaired Co-C bond rupture, as UV-visible spectroscopy did not show detectable AdoCbl homolysis upon binding of the physiological substrate, d-ornithine. AdoCbl homolysis was also not detected in the MCM mutants, as it was for the native enzyme. We conclude from these results that a gradual weakening of the electrostatic energy between the protein and the ribose leads to a progressive increase in the activation energy barrier for Co-C bond homolysis, thereby pointing to a key role for the conserved polar glutamate residue in controlling the initial generation of radical species.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Makins C,Pickering AV,Mariani C,Wolthers KRdoi
10.1021/bi3012719subject
Has Abstractpub_date
2013-02-05 00:00:00pages
878-88issue
5eissn
0006-2960issn
1520-4995journal_volume
52pub_type
杂志文章相关文献
BIOCHEMISTRY文献大全abstract::PT-ACRAMTU ([PtCl(en)(ACRAMTU-S)](NO(3))(2), en = ethane-1,2-diamine, ACRAMTU = 1-[2-(acridin-9-ylamino)ethyl]-1,3-dimethylthiourea) is a cytotoxic platinum-acridine conjugate previously shown to form adducts with the N3 endocyclic nitrogen of adenine in the DNA minor groove. This unusual observation and our prior det...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi050745n
更新日期:2005-08-23 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::Triton X-100 solubilized monomers of bacteiorhodopsin (bR) show a decrease in the extent of light adaptation; the red shift and the absorbance increase of the visible absorption band are reduced no less than half the values observed in purple membrane (p.m.) with a corresponding reduction in the isomerization of 13-ci...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00555a043
更新日期:1980-07-08 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::A metastable intermediate was found in the course of the denaturation of purified cytochrome P450 2B1 by increasing concentrations of guanidine hydrochloride (GuHCl). The metastable intermediate has no or low absorbance at 450 nm in the reduced carbon monoxide difference spectrum and has no absorbance at 420 nm. The i...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00016a023
更新日期:1995-04-25 00:00:00
abstract::Experimental measurements of the kinetic mechanism involving isoleucyl-tRNA synthetase proofreading valyl-tRNAIle in Escherichia coli have been incorporated into the conventional Michaelis-Menten model for this system. The model was subjected to a detailed mathematical analysis in the steady state. The results of this...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00303a019
更新日期:1984-04-10 00:00:00
abstract::The stoichiometry of cytochrome b559 (one or two copies) per reaction center of photosystem II (PSII) has been the subject of considerable debate. The molar ratio of cytochrome b559 has a number of significant implications on our understanding of the functional role of cytochrome b559, the mechanism of electron donati...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00161a024
更新日期:1992-11-24 00:00:00
abstract::Two new inhibitors in which the terminal α-carboxyl groups of Z-Ala-Ala-Phe-COOH and Z-Ala-Pro-Phe-COOH have been replaced with a proton to give Z-Ala-Ala-Phe-H and Z-Ala-Pro-Phe-H, respectively, have been synthesized. Using these inhibitors, we estimate that for α-chymotrypsin and subtilisin Carlsberg the terminal ca...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi300688k
更新日期:2012-08-07 00:00:00
abstract::Small molecule allostery modifies protein function but is not easily discovered. We introduce mass spectrometry integrated with equilibrium dialysis for the discovery of allostery systematically (MIDAS), a method for identifying physiologically relevant, low-affinity metabolite-protein interactions using unmodified pr...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi201313s
更新日期:2012-01-10 00:00:00
abstract::NSC 613862 (S)-(-) and NSC 613863 (R)-(+) are the two chiral isomers of ethyl-5-amino-2-methyl-1,2-dihydro-3-phenylpyrido[3, 4-b]pyrazin-7-yl carbamate. Both compounds bind to tubulin in a region that overlaps the colchicine site. They induce formation of abnormal polymers from purified GTP-Mg-tubulin, the active asse...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi970568t
更新日期:1998-01-13 00:00:00
abstract::BamA is an essential component of the β-barrel assembly machine (BAM) that is responsible for insertion and folding of β-barrel outer membrane proteins (OMPs) in Gram-negative bacteria. BamA is an OMP itself, and its β-barrel transmembrane domain is thought to catalyze OMP insertion and folding, although the molecular...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/acs.biochem.7b00281
更新日期:2017-06-20 00:00:00
abstract::The EcoRV DNA-(adenine-N6)-methyltransferase (MTase) recognizes GATATC sequences and modifies the first adenine residue within this site. Parts of its DNA interface show high sequence homology to DNA MTases of the dam family which recognize and modify GATC sequences. A phylogenetic analysis of M.EcoRV and dam-MTases s...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi0155450
更新日期:2001-09-18 00:00:00
abstract::The folate chromophore in native Escherichia coli DNA photolyase ([6R]-5,10-CH+-H4Pte(Glu)n=3-6) serves as an antenna, transferring light energy to the fully reduced flavin (FADH2) reaction center at high efficiency (EET = 0.92). Apophotolyase reconstituted after an overnight incubation with [6R,S]-5,10-CH+-H4folate (...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi9605548
更新日期:1996-06-18 00:00:00
abstract::A type II topoisomerase is essential for decatenating DNA replication products, and it accomplishes this task by passing one DNA duplex through a transient break in a second duplex. The B' domain of topoisomerase II contains three highly conserved motifs, EGDSA, PL(R/K)GK(I/L/M)LNVR, and IMTD(Q/A)DXD. We have investig...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi991328b
更新日期:2000-02-15 00:00:00
abstract::We have performed comparative studies of the neutral glycosphingolipids synthesized by three human myeloid leukemia cell lines, K562, KG1, and HL-60, which were metabolically labeled with [14C]galactose, to evaluate changes in neutral glycosphingolipid synthesis with myeloid cell differentiation. Individual neutral gl...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00345a034
更新日期:1985-11-19 00:00:00
abstract::The presence of high titers of anti-cardiolipin antibodies (ACA's) of autoimmune origin, which are known to bind to plasma beta2-glycoprotein I (aka apolipoprotein H), correlates clinically with autoimmune recurrent thrombosis. Soluble beta2-glycoprotein I binds to solid-phase ACA (immobilized on a surface plasmon res...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi9807207
更新日期:1998-11-17 00:00:00
abstract::Indolicidin is an antimicrobial cationic peptide with broad-spectrum activity isolated from bovine neutrophils. An indolicidin analogue CP-11, ILKKWPWWPWRRK-NH(2), with improved activity against Gram-negative bacteria had increased positive charge and amphipathicity while maintaining the short length of the parent mol...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi035643g
更新日期:2003-12-09 00:00:00
abstract::The Y70F mutant of aspartate aminotransferase has reduced affinity for coenzymes compared to the wild type. The equilibrium dissociation constants for pyridoxamine phosphate (PMP) holoenzymes, KPMPdiss, were determined from the association and dissociation rate constants to be 1.3 nM and 30 nM for the wild type and mu...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00244a014
更新日期:1991-07-30 00:00:00
abstract::Control of protein phosphatases is now understood to depend on binding to a variety of regulatory or targeting subunits to form holoenzymes with restricted localization and substrate specificity. In addition, the catalytic subunits of both type-1 and type-2 phosphatases bind specific inhibitor proteins. Here, we repor...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi992030o
更新日期:1999-12-21 00:00:00
abstract::Vitamin E is a dietary lipid that is essential for vertebrate health and fertility. The biological activity of vitamin E is thought to reflect its ability to quench oxygen- and carbon-based free radicals and thus to protect the organism from oxidative damage. However, recent reports suggest that vitamin E may also dis...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi701432q
更新日期:2008-01-15 00:00:00
abstract::The qualitative and quantitative features of mutagenesis by two DNA adducts of carcinogenic alkylating agents, O6-methylguanine (m6G) and O4-methylthymine (m4T), were examined in vivo. The deoxyhexanucleotides 5'-GCTAGC-3' and 5'-GCTAGC-3' were synthesized, where the underlined bases are the positions of m4T or m6G, r...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00242a031
更新日期:1991-07-16 00:00:00
abstract::Cytochrome P-450(M-1) [P-450(M-1)] is specifically expressed in adult male rat liver [Yoshioka, H., Morohashi, K., Sogawa, K., Miyata, T., Kawajiri, K., Hirose, T., Inayama, S., Fujii-Kuriyama, Y., & Omura, T. (1987) J. Biol. Chem. 262, 1706-1711]. Isolation and analysis of the gene for P-450(M-1) revealed that the co...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00399a039
更新日期:1987-12-15 00:00:00
abstract::Chromaffin granules isolated from bovine adrenal medulla were labeled with small unilamellar vesicles (SUV) made from N-(7-nitro-2,1,3-benzoxadiazol-4-yl)phosphatidylethanolamine and/or N-(lissamine rhodamine B sulfonyl)phosphatidylethanolamine as donor and acceptor fluorophores. Labeling of granules could be followed...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00315a020
更新日期:1984-09-25 00:00:00
abstract::4-Azido-2-nitrophenyl pyrophosphate (azido-PPi) labeled with 32P in the alpha position was prepared and used to photolabel beef heart mitochondrial F1. Azido-PPi was hydrolyzed by yeast inorganic pyrophosphatase, but not by mitochondrial F1-ATPase. Incubation of F1 with [alpha-32P]azido-PPi in the dark under condition...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00452a021
更新日期:1989-12-26 00:00:00
abstract::The three subunits I, II, and III of bovine procarboxypeptidase A separated by reversible dimethylmaleylation can reassociate to form the reconstituted complexes I + II, I + III, and I + II + III. Since the association II + III is not possible, subunit I appears to play a central role in the formation of the complex. ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00630a028
更新日期:1977-05-31 00:00:00
abstract::The functionality of the membrane-bound, ubiquinone-dependent pyruvate oxidase from the respiratory chain of Escherichia coli was reconstituted with a supported lipidic structure. The artificial structure was especially designed to allow the electrochemical control of the quinone pool through the lateral mobility of t...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi002325y
更新日期:2001-02-06 00:00:00
abstract::In this study, we prepared several shortened and full-length insulin analogues with substitutions at position B26. We compared the binding affinities of the analogues for rat adipose membranes with their ability to lower the plasma glucose level in nondiabetic Wistar rats in vivo after subcutaneous administration, and...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi702086w
更新日期:2008-05-27 00:00:00
abstract::Translocator protein 18 kDa (TSPO) was previously known as the peripheral benzodiazepine receptor (PBR) in eukaryotes, where it is mainly localized to the mitochondrial outer membrane. Considerable evidence indicates that it plays regulatory roles in steroidogenesis and apoptosis and is involved in various human disea...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/acs.biochem.6b00142
更新日期:2016-05-24 00:00:00
abstract::Apolipoprotein B (apoB) is the essential nonexchangeable protein in chylomicrons and very low-density lipoprotein-derived lipoprotein particles, including low-density lipoprotein (LDL). ApoB has been a key target for cardiovascular research because of its essential role in the assembly, secretion, delivery, and recept...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi048286y
更新日期:2005-02-01 00:00:00
abstract::The membrane-bound quinoprotein glucose dehydrogenase (mGDH) in Escherichia coli contains pyrroloquinoline quinone (PQQ) and participates in the direct oxidation of D-glucose to D-gluconate by transferring electrons to ubiquinone (UQ). To elucidate the mechanism of ubiquinone reduction by mGDH, we applied a pulse radi...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi051347n
更新日期:2005-10-18 00:00:00