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H A Dailey

Publications and source records attributed to H A Dailey.

81 records · Page 5Linked to original sources
PubMed1981

The role of COOH-terminal anionic residues in binding cytochrome b5 to phospholipid vesicles and biological membranes.

The COOH-terminal, anionic residues of the membrane binding segment of cytochrome b5 were examined to determine their possible significance in stabilizing the "tight" binding of the cytochrome to phospholipid vesicles. The removal of the 6 COOH-terminal residues, which include the carboxyl groups of Glu 132 and Asn 133, by carboxypeptidase digestion resulted in the loss of the characteristic "tight" binding to either synthetic phospholipid vesicles or isolated microsomes. Chemical modification of the four carboxyl groups of the nonpolar peptide of cytochrome b5 with carbodiimide and methylamine to produce a derivative with no anionic charged residues also resulted in a loss of this type of stable membrane interaction. These results suggest that the short polar COOH-terminal segment, containing two of the four carboxyl groups of the membrane binding domain of cytochrome b5, plays a crucial role in lipid-protein interactions that lead to the normal "tight" binding both in situ and in reconstituted phospholipid bilayer systems.

Amino Acid Sequence↗
PubMed1978

Structural and functional properties of the membrane binding segment of cytochrome b5.

Derivatives of cytochrome b5 that had been selectively shortened at the COOH-terminal, membrane binding segment of this amphipathic protein were employed to examine the minimum structural requirements for binding to phospholipid vesicles and for catalytic interactions in the stearyl-CoA desaturase system. Three derivatives shortened by 6, 18, and 27 amino acid residues were produced by controlled proteolysis with carboxypeptidases. The two largest derivatives bound to synthetic lipid vesicles and interacted with cytochrome b5. The third derivative neither bound to vesicles nor reacted with the desaturase. Whole nonpolar peptide and the nonpllar peptides of the two largest derivates contain only 29 to 34% polar residues, whereas the nonpolar peptide of the third derivative contains 44% polar residues. The secondary structure of the membrane binding segment was studied by circular dichroism of whole nonpolar peptide and the corresponding peptides of the three derivatives. The data for whole nonpolar peptide are consistent with a structure containing approximately 50% helical and 25% beta sheet structure. The CD of the nonpolar peptides of the two largest derivatives are consistent with structures containing 56% helix and 19% beta sheet structure, and 40% helix and 20% beta sheet structure. These data support a predicted model for secondary structure, proposed previously, based upon the primary structure (Fleming, P. J., Dailey, H. A., Corcoran D., and Strittmatter, P. (1978) J. Biol. Chem. 253, 5369-5372).

Amino Acid Sequence↗
PubMed1978

The primary structure of the nonpolar segment of bovine cytochrome b5.

The primary structure of the membrane bound segment of amphipathic bovine liver microsomal cytochrome b5 has been determined. This 43 residue nonpolar polypeptide is present at the COOH terminus of cytochrome b5. The sequence was obtained by automated sequence analysis and carboxypeptidase digestions. The sequence obtained is: Ile-Thr-Lys-Pro-Ser-Glu-Ser-Ile-Ile-Thr-Ile-Asp-Ser-Asn-Pro-Ser-Trp-Trp-Thr-Asn-Trp-Leu-Ile-Pro-Ala-Ile-Ser-Ala-Leu-Phe-Val-Ala-Leu-Ile-Tyr-His-Leu-Tyr-Thr-Ser-Glu-Asn. Conformational analysis using predictive algorithms is presented along with circular dichroism data on the peptide bound to phospholipid vesicles.

Amino Acid Sequence↗
PubMed1977

Reduction of iron and synthesis of protoheme by Spirillum itersonii and other organisms.

Membranes from Spirillum itersonii reduce ferric iron to ferrous iron with reduced nicotinamide adenine dinucleotide or succinate as a source of reductant. Iron reduction was measured spectrophotometrically at 562 nm using ferrozine, which chelates ferrous iron specifically. Reduced nicotinamide adenine dinucleotide or succinate was also effective as a source of iron. The effects of respiratory inhibitors suggested that reduction of iron occurs at one or more sites on the respiratory chain before cytochrome c. Reduction of iron and synthesis of protoheme with the physiological reductants were also observed with crude extracts of other bacteria, including Rhodopseudomonas spheroides, Rhodopseudomonas capsulata, Paracoccus denitrificans, and Escherichia coli. The effect of oxygen upon reduction of iron and formation of protoheme was examined with membranes from S. itersonii, using succinate as a source of reductant. Both systems were inhibited by oxygen, but this effect was completely reversed by addition of antimycin A. We conclude that reduced components of the respiratory chain serve as reductants for ferric iron, but with oxygen present they are oxidized preferentially by the successive members of the chain. This could be a mechanism for regulating synthesis of heme and cytochrome by oxygen.

Antimycin A↗
PubMed1977

Purification and characterization of the membrane-bound ferrochelatase from Spirillum itersonii.

The membrane-bound enzyme ferrochelatase (protoheme ferro-lyase, EC 4.99.1.1) was purified from isolated membrane fragments of Spirillum itersonii approximately 490-fold. Purification was achieved by solubilization with chaotropic salts followed by ammonium sulfate fractionation, diethylaminoethyl-cellulose chromatography, and gel filtration on Sephadex G-200. The purified enzyme has an apparent minimum molecular weight of approximately 50,000, as determined by gel filtration in the presence of 0.1% Brij 35 and 1 mM dithiothreitol but forms high-molecular-weight aggregates in the absence of detergent. Purified ferrochelatase is strongly stimulated in the presence of copper. The apparent Km for Fe2+ is 20 micrometer in the absence of copper and 9.5 micrometer in the presence of 20 micrometer CuCl2. The apparent Km for protoporphyrin is 50 micrometer, and it is unaltered by copper. Ferrochelatase has a single pH optimum of 7.50, and it is inhibited 50% by 20 micrometer heme. Certain divalent cations and sulfhydryl reagents also inhibit the enzyme.

Cell Membrane↗
PubMed1976

Membrane-bound respiratory of Spirillum itersonii.

The membrane-bound respiratory system of the gram-negative bacterium Spirillum itersonii was investigated. It contains cytochromes b (558), c (550), and o (558) and beta-dihydro-nicotinamide adenine dinucleotide (NADH) and succinate oxidase activities under all growth conditions. It is also capable of producing D-lactate and alpha-glycerophosphate dehydrogenases when grown with lactate or glycerol as sole carbon source. Membrane-bound malate dehydrogenase was not detectable under any conditions, although there is high activity of soluble nicotinamide adenine dinucleotide: malate dehydrogenase. When grown with oxygen as the sole terminal electron acceptor, approximately 60% of the total b-type cytochrome is present as cytochrome o, whereas only 40% is present as cytochrome o in cells grown with nitrate in the presence of oxygen. Both NADH and succinate oxidase are inhibited by azide, cyanide, antimycin A, and 2-n-heptyl-4-hydroxyquinoline-N-oxidase at low concentrations. The ability of these inhibitors to completely inhibit oxidase activity at low concentrations and their effects upon the aerobic steady-state reduction levels of b- and c-type cytochromes as well as the aerobic steady-state reduction levels obtained with NADH, succinate, and ascorbate-dichlorophenolindophenol suggest that presence of an unbranched respiratory chain in S. itersonii with the order ubiquinone leads to b leads to c leads to c leads to oxygen.

Antimycin A↗