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P K Warme

Publications and source records attributed to P K Warme.

At least 19 recordsLinked to original sources
PubMed1980

Semi-synthetic analogs of cytochrome c. Substitutions for methionine at position 80.

Derivatives of cytochrome c having S-methyl cysteine and ethionine substituted for methionine residue 80 have been synthesized in order to study the effects of structural perturbations near the heme on the biological function of cytochrome c. The etionine derivative has 96% as much activity as native cytochrome c in the succinate oxidase system, whereas the S-methyl cysteine analog is totally inactive.

Cyanogen Bromide↗
PubMed1979

Protected natural peptides as intermediates for preparing semisynthetic peptides and protein analogs. Selective acylation of epsilon-amino groups in cyanogen bromide peptides of cytochrome c using azidoformates.

Treatment of CNBr peptides 66--80, 81--104 and 66--104 from cytochrome c with t-butyloxycarbonyl azide leads to selective acylation of the epsilon-amino groups of lysine residues and the phenolic hydroxyl groups of tyrosine residues with less than 25% acylation of the alpha-amino groups. Similar selectivity was obtained for reactions of benzyloxycarbonyl azide, p-nitrobenzyloxycarbonyl azide and p-methoxybenzyloxycarbonyl azide with peptide 81--104. All of these protective groups can be removed under mild conditions, and thus, the partially protected natural peptides are desirable intermediates for the preparation of semisynthetic peptides. Model condensation reactions of N alpha t-butyloxycarbonyl methionine N-hydroxysuccinimide ester with Z-protected peptide 81--104 produced a peptide corresponding to residues 80 to 104 of cytochrome c in 94% yield.

Acylation↗
PubMed1979

Semisynthetic analogs of cytochrome c reconstructed from natural and synthetic peptides.

A biologically active semisynthetic hybrid of horse heart cytochrome c has been prepared by combining the heme peptide 1 through 65 (HP 1-65), prepared by CNBr cleavage of natural cytochrome c, with a semisynthetic peptide corresponding to positions 66 through 104. A fully protected synthetic peptide 66--79 was prepared by a modified solid phase peptide synthesis procedure and was converted to its N-hydroxysuccinimide ester. A peptide corresponding to residues 81--104 of cytochrome c was also isolated from the CNBr cleavage mixture and its epsilon-amino groups and tyrosyl hydroxyl group were protected selectively with the t-butyloxycarbonyl group. This partially protected peptide was reacted with t-butyloxycarbonyl methionine N-hydroxysuccinimide ester to give a derivative having methionine at position 80. This product was deprotected, purified and then t-butyloxycarbonyl groups were again introduced specifically on the epsilon-amino groups to give the peptide, Boc(Lys,Tyr)80--104. A semisynthetic peptide corresponding to residues 66 through 104 of cytochrome c was prepared by condensing the synthetic peptide 66--79 N-hydroxysuccinimide ester with t-butyloxycarbonyl (Lys,Tyr)80--104. The semisynthetic product was deprotected, purified and combined under anaerobic conditions with a heme peptide, HP 1-65, that was isolated from the products of CNBr cleavage of native cytochrome c. The reconstituted semisynthetic cytochrome c was purified by ion exchange chromatography and was shown to have the same oxygen uptake as native cytochrome c when assayed in the succinate oxidase system.

Acylation↗
PubMed1978

Chains of alternating sulfur and pi-bonded atoms in eight small proteins.

This paper demonstrates the existence of regions in eight small globular proteins in which the side chains of sulfur-containing amino acids (cysteine and methionine) alternate in space with side chains of aromatic amino acids (histidine, phenylalanine, tryptophan and tyrosine). The proteins are: rubredoxin, high potential iron protein, cytochrome c, flavodoxin, deoxyhemoglobin, trypsin inhibitor, ribonuclease-S, and lysozyme. The sulfur-pi-bonded 'chains' involve a minimum of five and a maximum of 10 amino acids, and contain the most polarizable atoms within proteins. S-pi-chains give extra stability to the folding of proteins; they may also afford paths for the step-wise movement of electrons.

Amino Acid Sequence↗
PubMed1977

Myoglobin as an oxygen indicator for measuring the oxygen binding characteristics of a modified myoglobin derivative containing covalently bound mesoheme.

By measuring the visible spectrum of a mixture of myoglobin and a modified derivative containing mesoheme in place of the normal protoheme, it is possible to evaluate the relative amounts of the oxidized, reduced, and oxygenated forms of each type of myoglobin. If the oxygen affinity of one myoglobin derivative is known, the oxygen affinity of the other can be determined from measurements at various oxygen partial pressures. In the absence of excess reducing agent, the rate of autoxidation can also be evaluated during the same experiment. The method described is suitable at very low oxygen partial pressures, where most previous methods are inaccurate, and it is very convenient to use, since no time-consuming calibration procedures are required. Using protoheme myoglobin as an oxygen indicator, the oxygen pressure at half saturation (P 1/2) of mesoheme myoglobin was shown to be 11% higher than the P 1/2 of a modified myoglobin derivative containing covalently bound mesoheme. The autoxidation rate of the covalent derivative is faster than that of the noncovalent derivative, but it is less dependent on oxygen pressure.

Chemical Phenomena↗
PubMed1975

Conformational energy refinement of horse-heart ferricytochrome c.

The reported X-ray structure of horse-heart ferricytochrome c has been refined by conformational energy calculations, using a three-stage computational procedure. In stage I, the atomic positions are adjusted to conform to idealized bond lengths and bond angles characteristic of small amino acid derivatives, while yet remaining as close as possible to the X-ray coordinates. In stage II, atomic overlaps are eliminated by adjusting the backbone and side-chain dihedral angles to minimize the nonbonded energy, hydrogen-bonded energy, and rotational energy contributions. In the final stage of refinement, the electrostatic energy and a more accurate hydrogen-bonded energy treatment are considered, in addition to the energy contributions of stage II. A "fitting potential" of gradually decreasing strength is imposed in both stages II and III, in order to keep the computed structure as similar to the x-ray structure as is consistent with a low-energy conformation. The final computed structure of cytochrome c exhibits a very low conformational energy (-504 kcal/mol) and also closely resembles the X-ray structure (RMS deviation = 0.77 A for all atoms). However, a special treatment was required in order to alter the location of the phenyl ring of phenylalanine-82. In contrast to the originally published X-ray structure, which shows the phenyl ring pointing away from the heme, the phenyl ring in the computed structure is tucked into the heme crevice, in a position similar to that observed in the reduced form of tuna cytochrome c, in the oxidized form of Rhodospirillum rubrum cytochrome c2, and also in the recently determined structure of oxidized tuna cytochrome c.

Animals↗
PubMed1975

The influence of amino acid substitutions on the conformational energy of cytochrome c.

Conformational energies have been evaluated for each of the staggered side-chain conformations associated with the 261 amino acid substitutions known to occur among 60 eucaryotic species. At least 86% of these substitutions can be sterically accommodated (one at a time) within the structure of horse-heart cytochrome c resulting from conformational energy refinement. Simultaneous incorporation of all pertinent amino acid substitutions found in eight representative species into the refined horse-heart structure is also shown to be sterically possible, with few exceptions. In two cases (Pekin duck cytochrome with 10 substitutions and Samia cynthia cytochrome with 24 substitutions), all substitutions could be readily incorporated, and the total energies associated with their computed structures differed by less than 10 kcal/mol from that of horse-heart cytochrome c. In the cytochromes from rattlesnake (22 substitutions), tuna (18 substitutions), and Neurospora crassa (36 substitutions), tyrosine could not be substituted for phenylalanine at position 46, within the constraints of the calculations. However, when all of the remaining substitutions were incorporated into these three cytochromes, their computed conformational energies differed by less than 30 kcal/mol from that of horse-heart cytochrome c. Between two and four amino acid substitutions cause high energies in the cytochromes from human, baker's yeast, and cotton seed, but all of the remaining substitutions are consistent with a low energy conformation. These results suggest that the structures of homologous proteins may be even more similar than has previously been recognized. Substitutions of all possible amino acid types at the invariant positions (where all eucaryotic cytochromes c bear the same amino acid) have revealed some cases where different amino acids can be accommodated, thus demonstrating that the biological constraints on amino acid substitutions are often different from the purely steric constraints investigated in this work.

Amino Acid Sequence↗
PubMed1974

Refinement of the X-ray Structure of Rubredoxin by Conformational Energy Calculations.

The x-ray structure of rubredoxin has been refined by energy minimization. The computed structure is constrained to have standard bond lengths, bond angles, and planar trans peptide groups. Also, since most of the steric overlaps have been relieved, it has a very low energy. As judged by the root mean square (RMS) deviation of the computed coordinates from those of the x-ray structure, the two are very similar. The reliability index R for the computed structure (determined from the structure factors for the calculated conformation) is 0.37, which is comparable to that for other proteins.

Journal Article↗
PubMed1973

Role of medium-range interactions in proteins.

The energies of oligopeptide segments of lysozyme are minimized with respect to the dihedral angles of the central residue. As the length of the oligopeptide segment increases, up to a nonapeptide, the low-energy conformation becomes that observed in the x-ray structure in most cases. This finding suggests that, while short-range interactions appear to play the dominant role in determining the conformation of an amino-acid residue in a protein, the additional interactions required to stabilize the conformation uniquely may be only of medium range, i.e., those within a nonapeptide, and longer-range interactions may be of considerably less importance.

Muramidase↗