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Guanidine hydrochloride denaturation helps determine the thermodynamic stability of a series of hydrophilic or hydrophobic surface mutants of iso-1-cytochrome c. These mutations lead to native state structural perturbations. Free energy of unfolding for these mutants are affected by both alpha-helix propensity and hydrophobic effects acting on the denatured state.

Magainin peptides are antimicrobial material that reduces the membrane activity of cytochrome oxidase liposomes. Magainin disrupts respiratory control and is proportional to lipid concentration. Respiratory control is revived to normal by proteolytic digestion of magainin. Magainin modulates multimeric pore that is responsible for membrane permeability and causes ion permeation that enhances respiratory activities.

Infrared spectroscopy facilitated the observation of an amide I infrared band shift which was influenced by guanidine hydrochloride concentration in the denatured state of a wild-type and two mutants of iso-1-cytochrome. The denatured-state infrared spectra was different from the wild-type protein spectra.

The effects of the deantured stateon the unfolding equilibrium of cytochrome c was investigated. Using site-directed mutagenesis, a series of lysine 73 iso-1-cytochrome c mutants were prepared by substitution with hydrophobic amino acids Met, Tyr, Phe and Trp. Lysine 73 is highly solvent-exposed and hydrophilic. The results indicate that the cytochrome c Lys73 mutants are more buried in the denatured than in the native state, that is, the protein in the denatured state adopts a nonrandom structure. Moreover, it was demonstrated that the mutations indeed significantly affected the folding equilibrium of cytochrome c.

Fourier transform spectroscopy was carried out to determine the secondary structure of hamster female protein (FP) in aqueous media and to evaluate calcium ion- and phosphorylcholine-induced conformational alteration. One half of the secondary structure of FP in the native form is in beta sheets with alpha helical and random structures in smaller proportion. Binding of calcium to the proposed binding sites at the regions spanning residue 93 to 109 and 150 to 168 alters FP's secondary structure. Further conformational changes is induces by phosphorylcholine. This conformational changes may explain the means by which FP's important functions are carried out.

Amide I infrared spectroscopy was used to determine the redox- and ionic strength-dependent changes in the secondary structure of cytochromes c from horse, cow, and dog hearts in water at 20 degrees Celsius. The beta-extended chain and turn structures exhibited sensitivity to the oxidation state of the heme iron. However, no significant structural alterations due to changes in ionic strength were determined. Moreover, it was demonstrated that amide I infrared spectroscopy is useful in the detection of subtle changes in the secondary structure within a protein.

Oxidation reactions of carbonyl species of hemoglobins A. Zurich, and Osier with ferricyanide to form met- hemoglobins have been investigated at pH 7.0 and 25 C. The kinetics support a two-step process in which, first, CO is lost to give the unliganded reduced species (deoxy), followed by oxidation in a one-electron transfer to ferricyanide ion. On and off rates for CO markedly affect the overall oxidation rate. That α subunits oxidize more rapidly than β subunits was noted by changes in C—O stretch frequency during the course of reactions with Hb A and Hb Zurich. The substitution of Arg for His at β63 in Hb Zurich resulted in a slower overall oxidation rate ascribable to greater affinity for CO. MetHb formation from stripped HbCO A was biphasic with the rate for the final ≈ 50% of the reaction about 30% faster titan that for the initial ≈ 50%. Addition of inositol hexakis- phosphate increased rates several-fold in both phases and shifted the point of change from slow to fast phase. These effects suggest that inositol hexakisphosphate binds weakly (Kd ≈ 30 μM) to Hb4(CO)4 in a high-affinity state which, after loss of one CO, changes to a low-affinity state where inositol hexakisphosphate binds more strongly (Kd < 1 μM) for the remainder of the reaction. With Hb Osler, which exhibits very low heme-heme interaction and reacts poorly with inositol hexakisphosphate, the reaction was monophasic with a rate throughout comparable to the initial rate for stripped HbCO A. The very low level of CO saturation (< 1–6%) usually found in vivo with normal hemoglobin subunits should not greatly affect overall rates of methemoglobin formation from one-electron electron acceptors. However, with the high levels of CO found bound (20–100% saturation) to the β subunit of hemoglobin Zurich in blood, the exposure to CO (as in smoking) can diminish the extent to which the abnormal Zurich subunit will be oxidized. CO exposure may. therefore, reduce significantly some of the undesired consequences of having hemoglobin Zurich in red blood cells. [ABSTRACT FROM AUTHOR]