Abstract:
:N-(1-Pyrenyl)iodoacetamide has been used to introduce fluorescent probes into Escherichia coli RNA polymerase. After an incubation time of 15 min, approximately 2 pyrene equiv was introduced per enzyme molecule. There was no further increase in modification after more extended periods of incubation. Neither calf thymus DNA nor nucleotides protected the holoenzyme from modification. Thus, the sites of modification do not appear to involve the binding sites for polynucleotides or the ribonucleoside triphosphates. From the isolation and analysis of the individual subunits, it was found that sigma contained approximately 1 pyrene equiv, beta contained 0.6, beta' contained 0.6, and alpha less than 0.1. Spectral and Stern-Volmer analyses indicate that the covalently attached pyrene molecules are in comparable apolar microenvironments. On the basis of CD analyses, the introduction of pyrene molecules into RNA polymerase alters its secondary structure. This alteration in secondary structure manifests itself by a reduction in overall enzymatic activity. Transcript analysis of the products obtained by using a linearized plasmid containing the A1 promoter and the Te terminator of bacteriophage T7 indicates that the pyrenyl derivative is capable of producing full-length transcripts and that it has an efficiency of chain termination comparable to the native enzyme. Analysis of tau plots for the interaction of the pyrenyl derivative and the native enzyme, respectively, with the A1 promoter yielded comparable values for the isomerization constant in the conversion of the closed complex to an open one. Comparable values were also obtained for the association constant. The rate of chain elongation for the pyrenyl derivative, however, is approximately 54% of that observed for the native enzyme. Thus, the decrease in overall transcriptional activity observed with the pyrenyl derivative is not due to a decrease in the efficiency of initiation or premature termination, but rather to a decrease in the rate of chain elongation.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Johnson RS,Bowers M,Eaton Qdoi
10.1021/bi00215a028subject
Has Abstractpub_date
1991-01-08 00:00:00pages
189-98issue
1eissn
0006-2960issn
1520-4995journal_volume
30pub_type
杂志文章相关文献
BIOCHEMISTRY文献大全abstract::Crystallographic studies of ligand-protein complexes reveal most preferable ligand binding modes, but do not show less populated modes that may contribute to measurable biochemical and biophysical characteristics of the complexes. In some cases, a ligand may bind a protein in essentially different modes. An example is...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi047553x
更新日期:2005-05-17 00:00:00
abstract::A new procedure for the synthesis of the pyrophosphate bond has been employed in the preparation of nucleoside dipyrophosphates from nucleoside 3',5'-diphosphates. The method makes use of a powerful phosphorylating agent generated in a mixture of cyanoethyl phosphate, dicyclohexylcarbodiimide, and mesitylenesulfonyl c...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00666a012
更新日期:1976-10-19 00:00:00
abstract::The interaction of profilin and non-muscle beta,gamma-actin prepared from bovine spleen has been investigated under physiologic ionic conditions. Profilin binding to actin decreases the affinity of actin for MgADP and MgATP by about 65- and 13-fold, respectively. Kinetic measurements indicate that profilin binding to ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi001520+
更新日期:2000-10-31 00:00:00
abstract::Carbamoyl phosphate synthetase from E. coli catalyzes the synthesis of carbamoyl phosphate through a series of four reactions occurring at three active sites connected by a molecular tunnel of 100 A. To understand the mechanism for coordination and synchronization among the active sites, the pre-steady-state time cour...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi992772h
更新日期:2000-05-02 00:00:00
abstract::Betaine-homocysteine S-methyltransferase (BHMT) is a zinc-dependent enzyme that catalyzes the transfer of a methyl group from glycine betaine (Bet) to homocysteine (Hcy) to form dimethylglycine (DMG) and methionine (Met). Previous studies in other laboratories have indicated that catalysis proceeds through the formati...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi049821x
更新日期:2004-05-11 00:00:00
abstract::Antithrombin III (ATIII) is a plasma-borne serine protease inhibitor that apparently forms covalent complexes with thrombin. The interaction between ATIII and thrombin is enhanced several thousandfold by the glycosaminoglycan, heparin. We have previously proposed that the heparin binding site of ATIII resides within a...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00490a010
更新日期:1990-09-25 00:00:00
abstract::13C NMR has been used to observe the equilibrium complex of [5,5-2H,5-13C]-5-aminolevulinate [( 5,5-2H,5-13C]ALA) bound to porphobilinogen (PBG) synthase (5-aminolevulinate dehydratase), a 280,000-dalton protein. [5,5-2H,5-13C]ALA (chemical shift 46.9 ppm in D2O) was prepared from [5-13C]ALA through enolization in deu...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00412a039
更新日期:1988-06-14 00:00:00
abstract::While two-component systems (TCSs), composed of a sensor histidine kinase (HK) and a response regulator, are the main signaling pathways in bacteria, global TCS activity remains poorly described. Here, we report the kinetic parameters of the HK autophosphorylation reaction using previously uncharacterized γ-phosphate-...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/acs.biochem.8b00485
更新日期:2018-07-24 00:00:00
abstract::Fibronectin is composed of multiple homologous repeats and contains many functional domains. Two major heparin-binding domains have previously been identified: the Hep I site near the amino terminus and the Hep II site near the carboxyl terminus. The Hep II site has been considered the high-affinity heparin-binding si...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi991624o
更新日期:2000-03-28 00:00:00
abstract::The type III secretion system (T3SS) is an essential virulence factor for Shigella flexneri , providing a conduit through which host-altering effectors are injected directly into a host cell to promote uptake. The type III secretion apparatus (T3SA) is composed of a basal body, external needle, and regulatory tip comp...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi400755f
更新日期:2013-12-10 00:00:00
abstract::Fully functional variable fragments (Fv) of D1.3, a mouse antibody directed against the hen egg lysozyme, were readily produced as hybrids (Fv-MalE) with the maltose-binding protein of Escherichia coli and purified independently of their antigen-binding properties. We used site-directed mutations of residues in the co...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi961419y
更新日期:1997-01-07 00:00:00
abstract::Previous optical and electron paramagnetic resonance (EPR) spectroscopic studies of the newly discovered peroxynitrite scavenging pseudoperoxidase from Leishmania major (LmPP) suggested that ferric LmPP contained a six-coordinate low-spin (6cLS) heme with a thiolate ligand, presumably a cysteine, bound to its heme iro...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi401343t
更新日期:2013-12-10 00:00:00
abstract::Myotoxin a from the venom of Crotalus viridis viridis (prairie rattlesnake) is a small protein which is responsible for myonecrosis. It is a basic protein with 42 amino acid residues of known sequence. Three disulfide bonds give it a highly compact structure. Microscopic examination of the toxin's effects reveals that...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00347a021
更新日期:1985-12-17 00:00:00
abstract::The distance between the calcium site (site I) and the zinc site (site II) in alpha-lactalbumin was estimated from Forster energy-transfer measurements between donor Eu(III) [or Tb(III)] at site I and acceptor Co(II) at site II to be 11.5 +/- 1.5 A. Intersite distances were also measured between the bis-ANS [4,4'-bis[...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00365a024
更新日期:1986-08-26 00:00:00
abstract::Ribonucleotides are frequently incorporated into DNA during replication. They are recognized and processed by several cellular enzymes, and their continued presence in the yeast nuclear genome results in replicative stress and genome instability. Thus, it is important to understand the effects of isolated ribonucleoti...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi201710q
更新日期:2012-03-27 00:00:00
abstract::Administration of pregnenolone-16 alpha-carbonitrile (PCN) to adult female rats caused a 2-fold increase in total liver microsomal cytochrome P-450 along with 5-7-fold increases in four in vitro monooxygenase activities considered diagnostic for the major PCN-inducible cytochrome P-450 isozyme. However, upon administr...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00387a022
更新日期:1987-06-30 00:00:00
abstract::A variant of the Tetrahymena ribozyme that efficiently cleaves single-stranded DNA under simulated physiological conditions [Tsang, J., & Joyce, G. F. (1994) Biochemistry 33, 5966-5973] was evaluated as a potential therapeutic agent on the basis of its ability to cleave synthetic oligonucleotide substrates correspondi...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi960845g
更新日期:1996-09-10 00:00:00
abstract::The interaction of aminooxy compounds such as aminooxyacetate (AAA), L-canaline, and hydroxylamine with sheep liver serine hydroxymethyltransferase (EC 2.1.2.1) was studied by absorption spectra and stopped-flow spectrophotometry and compared with the unique feature of interaction of O-amino-D-serine (OADS) with the e...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00451a010
更新日期:1989-12-12 00:00:00
abstract::We have described earlier a chromatin-bound protease with unique specificity for histone H2A [Eickbush, T. H., Watson, D. K., & Moudrianakis, E. N. (1976) Cell (Cambridge, Mass.) 9, 785--792]. In the present study, we explore the nature of interactions that form and stabilize the enzyme-chromatin system by using the a...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00531a008
更新日期:1982-01-19 00:00:00
abstract::Angiotensin converting enzyme (ACE) plays a critical role in the circulating or endocrine renin-angiotensin system (RAS) as well as the local regulation that exists in tissues such as the myocardium and skeletal muscle. Here we report the high-resolution crystal structures of testis ACE (tACE) in complex with the firs...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi049480n
更新日期:2004-07-13 00:00:00
abstract::Dihydroorotates deuteriated at both C5 and C6 have been prepared and used to probe the mechanism of the bovine liver mitochondrial dihydroorotate dehydrogenase. Primary deuterium isotope effects on kcat are observed with both (6RS)-[5(S)-2H]- and (6RS)-[6-2H] dihydroorotates (3 and 6, respectively); these effects are ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00429a041
更新日期:1989-02-07 00:00:00
abstract::Sedimentation in a preparative ultracentrifuge was used to determine the affinity of heavy meromyosin, HMM, for regulated actin, F-actin plus troponin-tropomyosin, in the presence of MgATP at pH 7.0, 20 degrees C, and mu = 18 mM. HMM was prepared from vertebrate striated muscle myosin by a mild chymotryptic digestion....
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00275a003
更新日期:1983-03-15 00:00:00
abstract::The N-terminal formamido group on imidazole- and pyrrole-containing polyamides causes stacked polyamides to bind in the minor groove of DNA in the staggered motif, and it also increases the binding affinity compared to those of non-formamido compounds. To further investigate the role of the N-terminal acylamido in aff...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi900242t
更新日期:2009-06-23 00:00:00
abstract::Previous crystallographic studies in this laboratory demonstrated that immunoglobulin light chains with the same amino acid sequence can have at least two and probably three or more conformations, depending on whether the second member of an interacting pair is a light or heavy chain. If a heavy chain is not available...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00594a022
更新日期:1978-01-10 00:00:00
abstract::Thymidylate synthase (TS) is a highly conserved homodimeric enzyme with two active sites, each of which contains amino acid residues from both subunits. We show that the conservation at the subunit interface between Escherichia coli TS and Lactobacillus casei TS is sufficient to permit the formation of a cross-species...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00090a002
更新日期:1993-10-05 00:00:00
abstract::The formation of amyloid fibril is associated with major human diseases, including Alzheimer's disease, prion diseases, and type 2 diabetes. Methods for efficient inhibition of amyloid fibril formation are therefore highly clinically important. A principal approach for the inhibition of amyloid formation is based on t...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi048582a
更新日期:2004-11-16 00:00:00
abstract::The rate of the reaction between p-nitroperoxybenzoic acid and cytochrome c peroxidase (CcP) has been investigated as a function of pH and ionic strength. The pH dependence of the reaction between CcP and peracetic acid has also been determined. The rate of the reactions are influenced by two heme-linked ionizations i...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi991497w
更新日期:1999-11-23 00:00:00
abstract::Two extremely potent inhibitors of citrate synthase, carboxyl and primary amide analogues of acetyl coenzyme A, have been synthesized. The ternary complexes of these inhibitors with oxaloacetate and citrate synthase have been crystallized and their structures analyzed at 1.70- and 1.65-A resolution, respectively. The ...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00191a002
更新日期:1994-06-28 00:00:00
abstract::Porcine parathyroid gland slices were incubated with 3H-labeled amino acids in order to label tissue proteins. After incubation a crude hormonal extract was prepared and analyzed by chromatography on carboxymethylcellulose. Among the three radioactive peaks which were detected in the eluate, two were identified as par...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00687a018
更新日期:1975-08-12 00:00:00
abstract::The direct electrochemistry of the cytochrome c/cytochrome b5 and cytochrome c/plastocyanin complexes has been investigated at edge-plane graphite and modified gold electrode surfaces, which are selective for one of the two components of the complex. Electrochemical response of one protein at an otherwise electrostati...
journal_title:Biochemistry
pub_type: 杂志文章
doi:10.1021/bi00465a010
更新日期:1990-04-03 00:00:00