The peripheral stalk participates in the yeast ATP synthase dimerization independently of e and g subunits.

It is now clearly established that dimerization of the F(1)F(o) ATP synthase takes place in the mitochondrial inner membrane. Interestingly, oligomerization of this enzyme seems to be involved in cristae morphogenesis. As they were able to form homodimers, subunits 4, e, and g have been proposed as potential ATP synthase dimerization subunits.

Failure to assemble the alpha 3 beta 3 subcomplex of the ATP synthase leads to accumulation of the alpha and beta subunits within inclusion bodies and the loss of mitochondrial cristae in Saccharomyces cerevisiae.

The F(1) component of mitochondrial ATP synthase is an oligomeric assembly of five different subunits, alpha, beta, gamma, delta, and epsilon. In terms of mass, the bulk of the structure ( approximately 90%) is provided by the alpha and beta subunits, which form an (alphabeta)(3) hexamer with adenine nucleotide binding sites at the alpha/beta interfaces. We report here ultrastructural and immunocytochemical analyses of yeast mutants that are unable to form the alpha(3)beta(3) oligomer, either because the alpha or the beta subunit is missing or because the cells are deficient for proteins that mediate F assembly (e.

The two rotor components of yeast mitochondrial ATP synthase are mechanically coupled by subunit delta.

The mitochondrial ATP synthase is made of a membrane-integrated F0 component that forms a proton-permeable pore through the inner membrane and a globular peripheral F1 domain where ATP is synthesized. The catalytic mechanism is thought to involve the rotation of a 10-12 c subunit ring in the F0 together with the gamma subunit of F1. An important and not yet resolved question is to define precisely how the gamma subunit is connected with the c-ring.

Topological and functional study of subunit h of the F1Fo ATP synthase complex in yeast Saccharomyces cerevisiae.

Subunit h, a 92-residue-long, hydrophilic, acidic protein, is a component of the yeast mitochondrial F1Fo ATP synthase. This subunit, homologous to the mammalian factor F6, is essential for the correct assembly and/or functioning of this enzyme since yeast cells lacking it are not able to grow on nonfermentable carbon sources. Chemical cross-links between subunit h and subunit 4 have previously been shown, suggesting that subunit h is a component of the peripheral stalk of the F1Fo ATP synthase.

Cellular localization of isoprenoid biosynthetic enzymes in Marchantia polymorpha. Uncovering a new role of oil bodies.

Like seed plants, liverworts synthesize and accumulate a myriad of isoprenoid compounds. Using antibodies raised against several isoprenoid biosynthetic enzymes, we investigated their intracellular compartmentation by in situ immunolocalization from Marchantia polymorpha. The enzymes examined were deoxy-xylulose phosphate synthase, geranyl diphosphate synthase, farnesyl diphosphate synthase, geranylgeranyl diphosphate synthase, monoterpene synthase, geranylgeranyl diphosphate reductase, phytoene synthase, and phytoene desaturase.

Investigation of the yeast mitochondrial unselective channel in intact and permeabilized spheroplasts.

The existence of an activity corresponding to the nucleotide-induced Yeast Mitochondria Unselective Channel (YMUC2) of isolated mitochondria was investigated in permeabilized and intact spheroplasts of the baker's yeast Yeast Foam. In nystatin-permeabilized spheroplasts, ATP and GDP-beta-S induced a decavanadate-sensitive stimulation of the respiration only under conditions equivalent to those previously reported for isolated mitochondria (low phosphate concentration, presence of a salt). On intact spheroplasts parallel measurements of respiration rate, [ATP]/[ADP] ratio and mitochondrial transmembrane potential allowed to show that the addition of the glucose analog 2-deoxyglucose decreased the permeability of the inner mitochondrial membrane owing to cellular ATP depletion.

Characterization of the Saccharomyces cerevisiae cytosine transporter using energizable plasma membrane vesicles.

The purine-cytosine permease is a carrier localized in the plasma membrane of the yeast Saccharomyces cerevisiae. The energetics of cytosine transport catalyzed by this permease has been studied in an artificial system obtained by fusion between proteoliposomes containing beef heart cytochrome c oxidase and plasma membrane-enriched fractions of a S. cerevisiae strain overexpressing the permease.

Modulation of the electrophoretic ATP-induced K(+)-transport in yeast mitochondria by delta pH.

The modulation of the ATP-induced K(+)-transport pathway of the yeast inner mitochondrial membrane by delta pH was investigated in two ways. First, the inhibitory effect of phosphate was compared to the effect of other permeant acids, demonstrating that a part of the effect of phosphate was linked to its electroneutral transport down delta pH. However, an additional effect specific for phosphate also occurred inside the matrix.

Functional analysis of mutated purine-cytosine permease from Saccharomyces cerevisiae. A possible role of the hydrophilic segment 371-377 in the active carrier conformation.

The purine-cytosine permease (PCP) is an active transporter located in the plasma membrane of the yeast Saccharomyces cerevisiae. This protein mediates purine (adenine, guanine, and hypoxanthine) and cytosine accumulation in the cell by using an electrochemical potential difference in proton as the energy source. Various mutant strains, with altered Kt(app) (apparent Michaelis constant of transport) of uptake for one or several bases, have already been selected.

Conditions allowing different states of ATP- and GDP-induced permeability in mitochondria from different strains of Saccharomyces cerevisiae.

The effect of ATP and other nucleotides on the respiration of Saccharomyces cerevisiae mitochondria was investigated. It was observed that ATP induced a stimulation of the respiration rate only in the presence of a salt in mitochondria from the baker's yeast Yeast Foam, whereas an ATP-induced stimulation occurred even in the absence of salt in mitochondria from three different laboratory strains. In both cases, the stimulation was related to a collapse of the transmembrane potential, suggesting the opening of ion- and/or proton-conducting pathways.

ATP synthase of yeast mitochondria. Isolation of the subunit h and disruption of the ATP14 gene.

A new subunit of the yeast ATP synthase (termed subunit h) has been isolated. Amino acid composition and N-terminal sequencing were determined by chemical methods. These data were in agreement with the sequence of the hypothetical protein L8003.

In vivo phosphorylation of the purine/cytosine permease from the plasma membrane of the yeast Saccharomyces cerevisiae.

The purine/cytosine permease, encoded by the FCY2 gene, is a carrier located in the plasma membrane of the yeast Saccharomyces cerevisiae. Polyclonal antibodies were raised against two peptides that corresponded to the sub-N-terminal and C-terminal sequences of the putative protein deduced from the FCY2 gene. Immunoprecipitation experiments performed with protein extracts labelled in vivo with 35S showed that purine/cytosine permease is specifically detected as a broad and diffuse band.

Stimulation of oxidative phosphorylation by electrophoretic K+ entry associated to electroneutral K+/H+ exchange in yeast mitochondria.

The effect of the addition of KCl, at constant osmolarity, was investigated on oxidative phosphorylation in isolated yeast mitochondria. KCl stimulated both respiration and ATP synthesis rates without changing the ATP/O ratio. KCl did not change the relationships between respiration rates and the protonmotive force.

Investigations of the inhibitory effect of propranolol, chlorpromazine, quinine, and dicyclohexylcarbodiimide on the swelling of yeast mitochondria in potassium acetate. Evidences for indirect effects mediated by the lipid phase.

The mode of action of propranolol, chlorpromazine, and quinine, three cationic drugs inhibiting swelling of yeast mitochondria in potassium acetate, was investigated by looking at their effect on fluorescent probes of the polar heads and of the nonpolar moiety of the membranes, under inhibitory conditions of swelling. As expected, propranolol and chlorpromazine exhibited specificity for anionic phospholipids since they increased the binding of the anionic probe 1-anilino 8-naphthalenesulfonate (ANS). Although propranolol did not release 1,6-diphenyl-1,3,5-hexatriene (DPH) from the hydrophobic moiety of the membrane, it increased the excimer/monomer fluorescence ratio of 10-(1-pyrene)decanoate, suggesting that it induced a limitation in the movements of the aliphatic chains of phospholipids.

Mutation in the hydrophobic domain of ATP synthase subunit 4 (subunit b) of yeast mitochondria disturbs coupling between proton translocation and catalysis.

We introduced mutations to test the function of the hydrophobic sector of subunit 4 from Saccharomyces cerevisiae ATP synthase. Mutations were introduced at the chromosomic locus by homologous transformation of a strain disrupted in the ATP4 gene. The strain carrying the replacement Leu68-Val69-->Arg-Glu did not grow at 37 degrees C owing to a lack of assembly of F1 and Fo sectors at this temperature.

Structural determination of bacterial nodulation factors involved in the Rhizobium meliloti-alfalfa symbiosis.

Extracellular signals produced by Rhizobium meliloti are able to induce root hair deformations and nodule organogenesis on alfalfa. The production of these signals is controlled by bacterial nod genes. To enable their isolation in significant amounts, an overproducting strain was constructed.

Isoniazid inhibition of mycolic acid synthesis by cell extracts of sensitive and resistant strains of Mycobacterium aurum.

Isonicotinic acid hydrazide (isoniazid; INH) inhibition of mycolic acid synthesis was studied by using cell extracts from both INH-sensitive and -resistant strains of Mycobacterium aurum. The cell extract of the INH-sensitive strain was inhibited by INH, while the preparation from the INH-resistant strain was not. This showed that the INH resistance of mycolic acid synthesis was not due to a difference in drug uptake or the level of peroxidase activity (similar in both extracts).

Minicell-forming mutants of Escherichia coli: suppression of both DicB- and MinD-dependent division inhibition by inactivation of the minC gene product.

We have determined the nucleotide sequence of the minB operon of 10 min mutants of Escherichia coli, characterized by impaired inhibition of polar divisions. These mutants were either sensitive or resistant to the division inhibitor DicB. All the mutations were found to lie in minC or minD, confirming the requirement of both gene products in the process of inhibition of polar sites.

Transposition in Shigella dysenteriae: isolation and analysis of IS911, a new member of the IS3 group of insertion sequences.

Twenty-nine clear-plaque mutants of bacteriophage lambda were isolated from a Shigella dysenteriae lysogen. Three were associated with insertions in the cI gene: two were due to insertion of IS600, and the third resulted from insertion of a new element, IS911. IS911 is 1,250 base pairs (bp) long, carries 27-bp imperfect terminal inverted repeats, and generates 3-bp duplications of the target DNA on insertion.

Structural and immunological properties of the phenolic glycolipids from Mycobacterium gastri and Mycobacterium kansasii.

Mycobacterial species-specific antigens belong to the three following classes: phenolic glycolipids (Phe Gl), acyltrehalose-containing lipooligosaccharides and polar glycopeptidolipids. These antigens have been chemically defined and alkali-labile epitopes were found to characterize the lipooligosaccharide antigen type. In the present study the major Mycobacterium kansasii phenolic glycolipid epitope namely Phe Gl K-I was delineated as the distal monoacetylated disaccharidic residue: 2,6-dideoxy-4-O-methyl-alpha-D-arabino-hexopyranosyl-(1----3)-2-O-methyl -4-O- acetyl-alpha-L-fucopyranose.

Evolution of large-subunit rRNA structure. The diversification of divergent D3 domain among major phylogenetic groups.

During evolution, the potential for sequence (and length) variation of large-subunit rRNA has been mostly restricted over 12 divergent domains (termed D1-D12) interspersed along the molecule. Here, we have focused our attention onto the D3 divergent domain, through a detailed analysis of its pattern of variation in the phylogeny, both in terms of primary and secondary structures. We have systematically compared all the procaryotic and eucaryotic sequences published so far (i.

Properties of new Escherichia coli Hfr strains constructed by integration of pSC101-derived conjugative plasmids.

Conjugative temperature-sensitive plasmids were derived from pSC101. These plasmids are useful in genetic analysis for two reasons: (i) they render possible the construction of new Hfr lines by plasmid integration at predetermined chromosomal loci via Tn10 inverse transposition, and (ii) the Hfr characters are transducible via bacteriophage P1. We also showed that replication from pSC101 origin is deleterious for the plasmid-chromosome fusion.

New-found phenolic glycolipids in Mycobacterium bovis BCG. Presence of a diglycosylated glycolipid.

A crude phenolic glycolipid extract from Mycobacterium bovis bacille Calmette-Guerin (BCG) was fractionated until homogeneity at the intact level into four phenolic glycolipids called B, B-1, B-2, and B-3 according to their polarity. The apolar one, which is the most abundant was assigned to the well-known mycoside B. The B-2 and B-3 phenolic glycolipids were purified by direct-phase high performance liquid chromatography using a 5 micron Spherisorb column but were only recovered in small amounts (3 mg).

Detection and possible role of two large nondivisible zones on the Escherichia coli chromosome.

Inversion of many predetermined segments of the Escherichia coli chromosome was attempted by using a system for in vivo selection of genomic rearrangements. Two types of constraints on these inversions were observed: (i) a sensitivity to rich medium when the distance between oriC and the 86- to 91-min region (which carries loci essential for transcription and translation) is increased; (ii) a poor viability or inviability of inversions having at least one endpoint in the one-third of the chromosome around replication terminators (with an exception for some inversions ending between these terminators). Although the first constraint is simply explained by a decreased dosage of the region involved, the second one may result from disruption of two long-range chromosomal organizations.

Tetraenoic and pentaenoic mycolic acids from Mycobacterium thamnopheos. Structure, taxonomic and biosynthetic implications.

On the basis of the analysis of mycolates, the type strain of Mycobacterium thamnopheos has been considered as a member of the genus Nocardia. In a comparative study conducted on mycobacterial species we found that M. thamnopheos synthesized two types of mycolate having the same mobilities on thin-layer chromatography as those of mycobacteria, but different from nocardomycolates.