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B Blazy

Publications and source records attributed to B Blazy.

5 recordsLinked to original sources
PubMed1979

Non-specific interactions of CRP from E. coli with native and denatured DNAs: control of binding by cAMP and cGMP and by cation concentration.

The cyclic adenosine 3',5'-monophosphate receptor protein of Escherichia coli (CRP) binds cooperatively to single- and double-stranded DNA. Binding data could be fitted to the model of McGhee and von Hippel (1) and show that neither strandedness of DNA, nor the effectors cAMP and cGMP or the ionic strength (KCl) do change appreciably the cooperativity parameter omega (omega approximately or equal to 100), and site size of DNA. Instead, distinctly different slopes were observed for the linear decrease of log K omega (a measure of the overall affinity) as a function of log (K+). From these double-log plots (2), the number of cations released and the non-electrostatic contributions to the binding free energy could be determined. Binding of CRP to single-stranded DNA is slightly favored under physiological ionic conditions (0.15-0.20 M), but such a preferential binding is almost abolished in the presence of cAMP which increases the strength of the interaction of the protein with both forms of DNA. CGMP does not change the binding properties and interactions of CRP with DNA. These observations do not support the proposal that the cAMP-CRP complex could stimulate transcription via some "melting" property unless its interactions be dramatically changed when it binds specifically to promoter DNA.

Bacteriophage lambda↗
PubMed1979

[Cooperative non-specific binding of the cyclic adenosine 3'--5'-monophosphate receptor protein (CRP) from Escherichia coli to double-stranded thymus and lambda pgal DNA].

Either free or combined with cAMP, CRP binds cooperatively to double-stranded thymus and lambda pgal DNA. The affinity of CRP for both DNAs in these non-specific interactions is increased by cAMP without noticeable change in the degree of cooperativity. Values of the intrinsic association constant, cooperativity parameter, and site size of DNA were determined from ultracentrifugal investigations under near-physiological ionic conditions.

Animals↗
PubMed1976

Mechanism of yeast cytochrome b2 action. I. Thermodynamics and relaxation kinetics of the interaction between cytochrome b2 and oxalate.

Oxalate is the strongest known inhibitor of yeast cytochrome b2 activity. We have used spectrophotometric titration, temperature-jump relaxation, and calorimetry in an investigation of the interaction between enzyme and inhibitor. The titration data are consistent with noncooperative binding to one site per subunit. This conclusion is corroborated by temperature-jump results which reveal a single relaxation phenomenon which obeys second-order kinetics. Further evidence for a simple binding reaction enthalpy estimated from relaxation amplitudes is in good agreement with the value obtained directly with batch calorimetry. The forward and reverse rate constants evaluated from the temperature-jump experiments are, respectively, 1 x 10(4) M-1 sec-1 and 15 sec-1. Although considerably smaller than a diffusion-controlled value, the forward rate constant is characterized by an unusually small activation energy of approximately 3 kcal/mol. This, together with a large unfavorable association activation entropy of -30 eu, suggests that oxalate diffuses freely to the active site, but only a small fraction of the collisions are productive due to severe steric requirements.

Binding Sites↗
PubMed1976

Mechanism of yeast cytochrome b2 action. II. Steady-state kinetics of oxalate inhibition.

From a careful steady-state kinetic study it is shown that the inhibition of L-lactate oxidation by cytochrome b2 with ferricyanide as acceptor is of the mixed competitive-noncompetitive type, indicating the formation of an active ternary complex between enzyme, substrate, and inhibitor. With a large excess of acceptor, the simplest formal mechanism consistent with all available data is: E + L equilibrium EL; E + S equilibrium ES leads to EP leads to E + P; ES + L equilibrium ESL leads to EPL leads to EL + P, where L is oxalate, S is L-lactate, P is pyruvate, and E is enzyme. The inhibition kinetics together with the rate constants for oxalate binding to free enzyme (Thusius, D., Blazy, B., and Baudras, A. (1976), Biochemistry, preceding paper in this issue) and recent steady-state experiments on L-lactate deuterated at C-2 (Lederer, F. (1974), Eur. J. Biochem, 46, 393) lead to estimates of some of the elementary rate parameters in the above scheme. As in the case of oxalate (see Thusius et al. reference above), the association rate constant for substrate binding (1.1 x 10(5) M-1 sec-1) is much smaller than a diffusion-controlled value. Our results also imply that dissociation of complex EP to free enzyme and pyruvate is partially rate limiting for the overall reaction.

Binding Sites↗