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Query: EC:3.6.3.14 (ATP synthase)
 7,042 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Protein synthesis was investigated in isolated mitochondria under conditions which either inhibited electron transport or uncoupled oxidative phosphorylation. In a medium containing an exogenous source of ATP and oligomycin, an inhibitor of the ATP synthase complex, incorporation of [35S]methionine into proteins is stimulated in the presence of inhibitors of the electron transport chain; substituting uncouplers of oxidative phosphorylation for the latter leads, in contrast, to a decrease in the rate of incorporation of the labeled amino acid into mitochondrial translation products. Studies on the metabolic stability of mitochondrial translation products revealed that "mature" polypeptides made in isolated mitochondria are stable as indicated by the absence of degradation during a 50 min "chase" period. Under conditions which reduce or dissipate the membrane potential, 50-60% of the newly made polypeptides (pulse) are degraded within 50 min. The kinetics of the degradation process for individual mitochondrial gene products reveal that the largest proportion of polypeptides degraded to an acid-soluble form during the chase period are abnormal proteins, likely the result of premature chain termination. Emerging as a common denominator in these studies is a role for a transmembrane potential across the inner membrane in the production of mature "stable" mitochondrial gene products.
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PMID:Expression of the mammalian mitochondrial genome. Role for membrane potential in the production of mature translation products. 233 40

The epitopes of two classes of monoclonal antibody and the binding site for the epsilon subunit have been mapped to the carboxyl-terminal region of the beta subunit of Escherichia coli F1-ATPase using partial CNBr cleavage, weak acid hydrolysis, and Western blots. One class of antibody, B-I, inhibits ATPase activity; the other class, B-II, recognizes an epitope not exposed on the surface of intact F1. Data from two-dimensional gels and blots of beta cleaved with CNBr/weak acid showed that the B-I epitope lies between Asp-381 and the carboxyl-terminal Leu-459, and the B-II epitope lies between Asp-345 and Met-380. Weak acid hydrolysis of the beta-epsilon product obtained by cross-linking F1 with a water-soluble carbodiimide yielded a fragment containing epsilon and a 13-kDa carboxyl-terminal fragment of beta indicating that epsilon interacts with this portion of beta as well. Fab fragments from the B-I antibody beta-6 could be cross-linked to the epsilon subunit in native F1 by various cross-linking agents demonstrating that the antibody and the epsilon subunit occupy adjacent, nonoverlapping sites on the beta subunit. Implications of these results for the roles of the epsilon subunit and of the carboxyl-terminal region of the beta subunit in F1 are discussed.
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PMID:The epsilon subunit and inhibitory monoclonal antibodies interact with the carboxyl-terminal region of the beta subunit of Escherichia coli F1-ATPase. 244 Aug 72

Three monoclonal antibodies (mAbs) recognizing distinct epitopes on the delta-subunit of beef heart mitochondrial F1-ATPase were studied for their reactivity towards the delta-subunit both in isolated F1 and in the F0-F1 complex of submitochondrial particles. Two of the antibodies termed mAb delta 195 and mAb delta 239 had free access to delta in F1 and the F0-F1 complex. Partial hindrance was observed for the third antibody mAb delta 22. By a double antibinding assay, it was found that the binding sites for mAb delta 195 and mAb delta 239 were close to each other and possibly overlapping. Mapping studies conducted with the isolated delta-subunit showed that mAb delta 195 and mAb delta 239 interacted with the N-terminal portion of delta extending from Ala-1 to Met-16, whereas mAb delta 22 interacted with the fragment spanning Ser-17-Glu-68. It was concluded that the Ala-1-Met-16 segment of the delta-subunit in F1 and the F0-F1 complex is freely accessible from the outside, whereas the Ser-17-Glu-68 segment of delta is partially hidden, possibly as a result of interactions with other subunits.
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PMID:Exploration of delta-subunit interactions in beef heart mitochondrial F1-ATPase by monoclonal antibodies. 246 98

A site-directed mutation in the gene which codes for the c-subunit of the F1F0-ATPase, resulting in the substitution of Ala-25 by Tyr, has been constructed and characterized. A plasmid carrying the mutation was used to transform strain AN943 (uncE429). The resulting strain is unable to grow on succinate as sole carbon source and possesses an uncoupled growth yield. Membranes prepared from the mutant possess low levels of ATPase activity and are proton-impermeable. The F1-ATPase activity was found to be inhibited by 80% when bound to the membrane. When carried on a plasmid, the mutation is dominant in complementation tests with all mutant unc alleles tested and when transformed into wild-type strain AN346, the mutation results in an uncoupled phenotype. A mutant which overcomes this dominance was isolated and found to possess an 11-amino-acid deletion extending from Ile-55 to Met-65 within the c-subunit. These results are discussed in relation to the previously isolated Ala-25 to Thr mutant (Fimmel, A.L., Jans, D.A., Hatch, L., James, L.B., Gibson, F. and Cox, G.B. (1985) Biochim. Biophys. Acta 808, 252-258) and in relation to a previously proposed model for the F0 (Cox, G.B., Fimmel, A.L., Gibson, F. and Hatch, L. (1986) Biochim. Biophys. Acta 849, 62-69).
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PMID:The F1F0-ATPase of Escherichia coli. The substitution of alanine by tyrosine at position 25 in the c-subunit affects function but not assembly. 252 60

A gene was discovered in the cyanobacterium Synechococcus 6301 that encodes a protein highly related to members of the [2Fe-2S] ferredoxin family found in chloroplasts and cyanobacteria. It follows a cluster of seven genes encoding subunits of the cyanobacterial ATP synthase complex. It is transcribed as a monocistronic mRNA of 408 nucleotide residues. Transcription starts at a site 55 bp upstream of the initiator methionine codon. Transcriptional initiation and termination signals with sequences similar to those found in Escherichia coli are not present. Comparison of the predicted sequence of the ferredoxin protein with those of other cyanobacterial and plant ferredoxins shows an average sequences identity of about 40%. Twelve amino acid residues are invariant, including the four cysteine residues that provide ligands for the [2Fe-2S] cluster. The deduced Synechococcus ferredoxin protein sequence has a C-terminal extension of eight amino acid residues relative to most other 2Fe-2S ferredoxins except for those from halobacteria, which also have a C-terminal extension. The sequence of the Synechococcus protein is most closely related to ferredoxins from the two complex cyanobacteria Chlorogloeopsis fritschii and Mastigocladus laminosus. The deduced protein sequence is not that of the major soluble ferredoxin that has been isolated from Synechococcus 6301 and is reported in the accompanying paper [Wada, Masui, Matsubara & Rogers (1988) Biochem. J. 252, 571-575]. So it appears to be a novel [2Fe-2S] ferredoxin and Synechococcus 6301 contains at least two [2Fe-2S] ferredoxins, which may have different roles in vivo.
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PMID:Expression of a gene encoding a novel ferredoxin in the cyanobacterium Synechococcus 6301. 284 73

The nucleotide sequence of the yeast mitochondrial olil gene has been obtained in a series of mit- mutants with mutations in this gene, which codes for subunit 9 of of the mitochondrial ATPase complex. Subunit 9 is the proteolipid, 76 amino acids in length, necessary for the proton translocation function of the membrane Fo-sector. These mutants were classified on the basis of their rescue by a petite strain shown here to retain the entire wild-type olil gene. The mutation in one mit- strain removes a positively charged residue (Arg39----Met) which is likely to be located in a segment of subunit 9 that protrudes from the inner mitochondrial membrane. In a second mit- mutant, a negatively charged residue replaces a conserved glycine residue (Gly18----Asp) in a glycine-rich segment of the protein that is most likely embedded within the membrane. Other mit- mutations result in frameshifts with predicted products 7, 65 and 68 amino acid residues long. In each mit- mutant, there is the loss of one or more of the amino acid residues that are highly conserved among diverse species. The location and nature of specific changes pinpoint amino acid residues in subunit 9 essential to the activity of the mitochondrial ATPase complex.
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PMID:Biogenesis of mitochondria: DNA sequence analysis of mit- mutations in the mitochondrial oli1 gene coding for mitochondrial ATPase subunit 9 in Saccharomyces cerevisiae. 286 Jun 38

When beef heart mitochondrial F1-ATPase is photoirradiated in the presence of 2-azido[alpha-32P]adenosine diphosphate, the beta subunit of the enzyme is preferentially photolabeled [Dalbon, P., Boulay, F., & Vignais, P. V. (1985) FEBS Lett. 180, 212-218]. The site of photolabeling of the beta subunit has been explored. After cyanogen bromide cleavage of the photolabeled beta subunit, only the peptide fragment extending from Gln-293 to Met-358 was found to be labeled. This peptide was isolated and digested by trypsin or Staphylococcus aureus V8 protease. Digestion by trypsin yielded four peptides, one of which spanned residues Ala-338-Arg-356 and contained all the bound radioactivity. When trypsin was replaced by V8 protease, a single peptide spanning residues Leu-342-Met-358 was labeled. Edman degradation of the two labeled peptides showed that radioactivity was localized on the following four amino acids: Leu-342, Ile-344, Tyr-345, and Pro-346.
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PMID:Identification of amino acid residues photolabeled with 2-azido[alpha-32P]adenosine diphosphate in the beta subunit of beef heart mitochondrial F1-ATPase. 287 32

One subunit of the membrane portion of yeast ATP synthase was purified. Structural data are reported. This subunit (subunit 4) is the fourth polypeptide of the complex when classifying subunits in order of decreasing molecular mass. Its apparent relative molecular mass is about 25,000. The polypeptide was extracted from the complex with a mixture of chloroform/methanol (1/1) and 0.5 M pyridinium acetate pH 6.0. Purification was performed with a combination of gel permeation chromatography on Sephadex G-75 and high-performance gel permeation chromatography with aqueous solvents containing 5% sodium dodecyl sulfate. The amino acid composition is reported here. The following sequence of the NH2-terminal ten residues was determined: Met-Ser-Ser-Thr-Pro-Glu-Lys-Gln-Thr-Asp.
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PMID:Subunit 4 of ATP synthase (F0F1) from yeast mitochondria. Purification, amino-acid composition and partial N-terminal sequence. 288 7

Six mutant uncD alleles, affecting essential residues of the beta-subunit of Escherichia coli proton-ATPase, have been identified by intragenic complementation mapping, cloning, and DNA sequencing. Five of the mutations impair catalysis but do not cause structural perturbation of F1-ATPase. The amino acid substitutions found were as follows: uncD412, Gly-142----Ser; uncD430 and uncD431, both Arg-246----Cys; uncD478, Ser-174----Phe; and uncD484, Met-209----Ile. Kinetic characteristics of each corresponding mutant F1-ATPase are described or reviewed. In each case, the major determinant of impaired catalysis appears to be an attenuation of positive catalytic site cooperativity. Additionally, each mutation affects intrinsic properties of the catalytic site, including affinity for ATP, the ratio between unisite-bound substrate and products, and the rate of release of product inorganic phosphate under unisite ATP hydrolysis conditions. These effects are discussed in terms of a structural model of the catalytic nucleotide-binding domain of beta-subunit proposed recently (Duncan, T.M., Parsonage, D., and Senior, A.E. (1986) FEBS Lett. 208, 1-6). Each of the mutations lies within that domain. The uncD409 allele abolishes normal assembly of F1-ATPase. The amino acid substitution is Gly-214----Arg, which is suggested to affect a beta-turn connecting a beta-strand and an alpha-helix in the predicted nucleotide-binding domain of the beta-subunit.
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PMID:The defective proton-ATPase of uncD mutants of Escherichia coli. Identification by DNA sequencing of residues in the beta-subunit which are essential for catalysis or normal assembly. 288 84

Bovine cardiac muscle was extracted by an acidic chloroform/methanol mixture. A combination of gel permeation and ion-exchange chromatographies in organic solvents and HPLC allowed the purification of subunits VIIIa (Mr 5400) and VIIIb (Mr 4900) of cytochrome c oxidase and of A6L protein (Mr 7900) of ATP synthase. The identification of the proteins was made possible by measurement of their molecular weight by fast atom bombardment-mass spectrometry (FAB-MS) in conjunction with conventional Edman degradation. The determination by FAB-MS of the molecular weight of A6L protein confirmed its supposed formylated N-terminal methionine.
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PMID:Purification and characterization of low-molecular-weight beef heart proteolipids: use of fast atom bombardment-mass spectrometry for identification. 290 96


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