Mechanism of an insect glutathione S-transferase: kinetic analysis supporting a rapid equilibrium random sequential mechanism with housefly I1 isoform.

The steady-state kinetics of glutathione S-transferase I1 (GST I1) from housefly Musca domestica expressed in Escherichia coli were investigated with glutathione (GSH) and 1-chloro-2,4-dinitrobenzene (CDNB). Concentrations of the varied substrates were from 0.03 to 1 mM for GSH and 0.

Near ultraviolet circular dichroism study of the cyclic AMP receptor protein, its NH2-terminal domain and their interaction with cyclic AMP.

Circular dichroism in the near ultraviolet wavelength range was employed to examine conformational features of CRP (a dimer with a chain of 209 amino acids) and of its subtilisin core -alpha CRP- which retains the cAMP binding site (a dimer spanning the sequence 1-117). Binding of the ligand cAMP (allosteric activator), as well as cGMP was also investigated. The well resolved transitions could be assigned to the various classes of aromatic amino acid residues in the two proteins.

Ligand-modulated binding of a gene regulatory protein to DNA. Quantitative analysis of cyclic-AMP induced binding of CRP from Escherichia coli to non-specific and specific DNA targets.

This paper describes a generally applicable method for quantitative investigation of ligand-dependent binding of a regulatory protein to its target DNA at equilibrium. It is used here to analyse the coupled binding equilibria of cAMP receptor protein from Escherichia coli K12 (CRP) with DNA and the physiological effector cAMP. In principle, the DNA binding parameters of CRP dimers with either one or two ligands bound are determinable in such an approach.

On the origin of selectivity in recognition by cyclic adenosine 3',5'-monophosphate receptor protein of its specific binding site of the lactose promoter region.

From fluorescence measurements we could analyse the binding of cyclic adenosine 3',5'-monophosphate receptor protein (CRP) from Escherichia coli to its specific site on a 301 base-pair long DNA fragment containing the control region of the lactose operon. At physiological ionic strength selection of the specific site is strictly dependent on the allosteric effector cAMP, and binding of the cAMP . CRP complex to its specific site is favoured over the non-specific binding by 5 kcal/mol with Kass (specific) = 10(8) M-1 at 37 degrees C.

[Specific association of the CRP-cyclic AMP complex with the control region of the lactose operon of Escherichia coli K 12: a direct fluorimetric study using DNA fragments of different lengths].

Specific-site binding of the cAMP . CRP complex to the control region of the lactose operon of E. coli was measured directly.