Michael H. Klapper

On the nature of the protein interior

Abstract An estimate of the atomic densities of six proteins suggests that the protein interior is very densely packed, and that the ‘oil-drop’ picture is not an adequate model. On this basis the compressibility change accompanying protein denaturation is predicted to be positive, as is the volume change on chain unfolding. These predictions are consistent with experimental observations.

The independent distribution of amino acid near neighbor pairs into polypeptides

Abstract With the assumption that individual amino acids are independently distributed into naturally occurring polypeptide chains, it is shown that amino acid pairs with 0–2 arbitrary intervening residues are also independently distributed, with a few possible exceptions. This is not true of N- and C-terminal amino acids.

Long range electron transfer between tyrosine and tryptophan in hen egg-white lysozyme

The azide, dibromide and dichloride radicals oxidize one or more tryptophan side chains in hen egg-white lysozyme. The indolyl radical produced in this second-order 1-electron oxidation subsequently oxidizes a tyrosine side chain to the phenoxy radical in an intramolecular reaction with a rate constant of 130 +/- 10 s-1 at pH 7, 25 degrees C. The final indolyl and phenoxy equilibrium mixture then decays with a t1/2 approximately 2 s. The faster intramolecular reaction exhibits a pH dependence; on decreasing the pH from 9 the first-order rate constant increases to a maximum near pH 5.4 and then declines as the pH is lowered further. In contrast, the first-order rate constant for the intramolecular electron transfer between the tyrosine and tryptophan of the peptide trpH-pro-tyrOH remains unchanged between approx. pH 11 and 6.5 and then increases as the pH is lowered further. This difference in the observed pH dependence suggests that changes in structure or ionization state influence the protein electron transfer rate. We also discuss the radiation inactivation of lysozyme in light of these observations.

Long range electron transfer along an α-helix

Abstract The many observations of long range electron transfer in proteins raises the question of whether a proteins structure can influence the rate or path of such transfers, and if so, then how. To answer these questions requires information on which of the various structural elements composing proteins support long range electron transfer. In this report, we present evidence for long range electron transfer along the α-helix of a synthetic leucine zipper dimer. We also present electron transfer rate data obtained with other helical peptides.

Rapid binding of PMB to non-SH groups on hemerythrin☆

Abstract Using the flow dialysis technique we find that p-mercuribenzoate binds to the non-heme iron protein hemerythrin at a number of sites other than the sulfhydryl group. This non-sulfhydryl binding probably occurs faster than formation of the sulfur-mercury bond.

The ‘knotting’ problem in proteins. Loop penetration

The knotting problem in proteins is redefined in terms of the more general concept of loop penetration. Disulfide-containing proteins are represented as linear graphs, and it is proposed that proteins with nonplanar graph representations display loop penetrations in their three-dimensional structure. With this argument, it is shown that the occurrence of loop penetration in disulfide-containing proteins is approximately that expected based on the random pairing of sulfhydryl groups. It is argued that loop penetration is not as rare as currently held.

Catalytic activity of Nτ-Carboxymethylhistidine-12 ribonuclease: pH dependence

The pH-dependence of RNAase A and of Ntau-carboxymethylhistidine-12-RNAase (ribonucleate 3-pyrimidino-oligonucleotidohydrolase) catalysis was studied. Apparent acid dissociation constants were obtained by least squares analysis of the kinetics data. These dissociation constants were compared with pKa values of model imidazole compounds, and with pKa values of histidine residues 12 and 119 on the protein. The shapes of the kcat versus pH profiles for RNAase A and its carboxymethyl derivative are very similar, from which it is concluded that the mechanism of catalysis is closely similar in the two proteins. Apparent pKa values obtained from the kinetic data are higher for the carboxymethylated protein than for RNAase A, as are the pKa values of residues 12 and 119. The similar shifts are consistent with the conclusions that both these residues are functionally significant in native and modified enzyme, and that an unblocked tau-nitrogen on histidine-12 is not essential for activity. From the enzymes catalytic dependence on pH, and the NMR determined pKa values we propose that histidine 12 and 119 function catalytically in their basic and acidic forms respectively.

The methoxatin semiquinone: A pulse radiolysis study

Methoxatin is a novel o-quinone found in bacterial dehydrogenases and mammalian plasma amine oxidase. This is the first report of the redox potential and spectrum of the 1-electron reduced methoxatin semiquinone obtained by the method of pulse radiolysis in aqueous solution.

Pseudo-dynamic contact surface areas: estimation of apolar bonding

A new Monte Carlo based algorithm has been written for the computation of pseudo-dynamic contact surface areas. The linear correlation of this contact area with solute transfer free energies (water leads to organic liquid) is established for apolar amino acid side chains. The slope of these linear plots, deltaGosp, is a unitary free energy which has potential use in the estimation of apolar bond free energies in proteins. The magnitude of deltaGosp is dependent upon the nature of the organic solvent involved in the transfer process, varying from 86 to 130 cal/A2. Analogues linear correlations with the same range of deltaGosp values are observed for inhibitors of protein-catalyzed reactions.

Sequential measurements of glycolytic and tricarboxylic acid cycle metabolites in single sample aliquots.

Abstract Standard, pyridine nucleotide-linked fluorometric assays have been sequenced so that 14 glycolytic and tricarboxylic acid cycle intermediates can be measured in only two sample aliquots. This multiple assay procedure, which is based on commercially available enzymes and reagents, allows analysis on small tissue samples in relatively short times.