Journal of Bacteriology

Seiboth, B; Hakola, S; Mach, RL; Suominen, PL; Kubicek, CP

doi: N/Apmid: 9286982

B Seiboth, S Hakola, RL Mach, PL Suominen and CP Kubicek Abteilung fur Mikrobielle Biochemie, Institut fur Biochemische Technologie und Mikrobiologie, TU Wien, Vienna, Austria. The relative contributions of four major cellulases of Trichoderma reesei (1,4-beta-D-glucan cellobiohydrolase I CBH I, CBH II, endo-1,4- beta-D-glucanase I EG I, and EG II) to the generation of the cellulase inducer from cellulose were studied with isogenic strains in which the corresponding genes (cbh1, cbh2, egl1, and egl2) had been deleted by insertion of the Aspergillus nidulans amdS marker gene. During growth on lactose (a soluble carbon source provoking cellulase gene expression), these strains showed no significant alterations in their ability to express the respective other cellulase genes, with the exception of the strain containing delta cbh1, which exhibited an increased steady-state level of cbh2 mRNA. On crystalline cellulose as the only carbon source, however, significant differences were apparent: strains in which cbh2 and egl2, respectively, had been deleted showed no expression of the other cellulase genes, whereas strains carrying the cbh1 or egl1 deletion showed these transcripts. The delta cbh1- containing strain also showed enhanced cbh2 mRNA levels under these conditions. A strain in which both cbh1 and cbh2 had been deleted, however, was unable to initiate growth on cellulose. Addition of 2 mM sophorose, a putative inducer of cellulase gene expression, to such cultures induced the transcription of egl1 and egl2 and restored the ability to grow on cellulose. We conclude that CBH II and EG II are of major importance for the efficient formation of the inducer from cellulose in T. reesei and that removal of both cellobiohydrolases renders T. reesei unable to attack crystalline cellulose.

Thorstenson, Y R; Zhang, Y; Olson, P S; Mascarenhas, D

doi: N/Apmid: 9286985

Leaderless polypeptides efficiently extracted from whole cells by osmotic shock. Y R Thorstenson , Y Zhang , P S Olson and D Mascarenhas Department of Molecular & Cell Biology, Celtrix Pharmaceuticals, Santa Clara, California 95054, USA. ABSTRACT Three molecular foldases, DsbA, DsbC, and rotamase (ppiA), exhibited the unusual property of accumulating in an osmotically sensitive cellular compartment of Escherichia coli when their signal sequences were precisely removed by mutation. A mammalian protein, interleukin-1 (IL-1) receptor antagonist, behaved in a similar fashion in E. coli when its native signal sequence was deleted. These leaderless mutants (but not two control proteins overexpressed in the same system) were quantitatively extractable from whole cells by a variety of methods generally employed in the recovery of periplasmic proteins. A series of biochemical and genetic experiments showed that (i) leaderless DsbA (but not the wild type) was retained in a nonperiplasmic location; (ii) beta-galactosidase fusions to leaderless DsbA (but not to the wild type) exhibited efficient alpha complementation; (iii) none of the leaderless mutant proteins were substantially associated with cell membranes, even when they were overexpressed in cells; and (iv) leaderless DsbA was not transported to an osmotically sensitive compartment via a secA- or ftsZ-dependent mechanism. The observation that these proteins transit to some privileged cellular location by a previously undescribed mechanism(s)--absent their normal mode of (signal sequence-dependent) translocation--was unexpected. DsbA, rotamase, and IL-1, whose tertiary structures are known, appear to be structurally unrelated proteins. Despite a lack of obvious homologies, these proteins apparently have a common mechanism for intracellular localization. As this (putative) bacterial mechanism efficiently recognizes proteins of mammalian origin, it must be well conserved across evolutionary boundaries. CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? « Previous | Next Article » Table of Contents This Article J. Bacteriol. September 1997 vol. 179 no. 17 5333-5339 » Abstract PDF Services Email this article to a colleague Similar articles in ASM journals Alert me when this article is cited Alert me if a correction is posted Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of JB Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Load citing article information Citing articles via Web of Science Citing articles via Google Scholar Google Scholar Articles by Thorstenson, Y. R. Articles by Mascarenhas, D. Search for related content PubMed PubMed citation Articles by Thorstenson, Y. R. Articles by Mascarenhas, D. Related Content Load related web page information Social Bookmarking CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? current issue December 2011, volume 193, issue 24 Alert me to new issues of JB About JB Subscribers Authors Reviewers Advertisers Inquiries from the Press Permissions & Commercial Reprints ASM Journals Public Access Policy JB RSS Feeds 1752 N Street N.W. • Washington DC 20036 202.737.3600 • 202.942.9355 fax • [email protected] Print ISSN: 0021-9193 Online ISSN: 1098-5530 Copyright © 2011 by the American Society for Microbiology. For an alternate route to JB .asm.org, visit: http://intl- JB .asm.org | More Info»

Miranda, M; Ramos, A; Veiga-da-Cunha, M; Loureiro-Dias, MC; Santos, H

doi: N/Apmid: 9286987

M Miranda, A Ramos, M Veiga-da-Cunha, MC Loureiro-Dias and H Santos Instituto de Tecnologia Quimica e Biologica/Instituto de Biologia Experimental e Tecnologica, Universidade Nova de Lisboa, Oeiras, Portugal. The sugar-induced inhibition of malolactic fermentation in cell suspensions of Leuconostoc oenos, recently reclassified as Oenococcus oeni (L. M. T. Dicks, F. Dellaglio, and M. D. Collins, Int. J. Syst. Bacteriol. 45:395-397, 1995) was investigated by in vivo and in vitro nuclear magnetic resonance (NMR) spectroscopy and manometric techniques. At 2 mM, glucose inhibited malolactic fermentation by 50%, and at 5 mM or higher it caused a maximum inhibitory effect of ca. 70%. Galactose, trehalose, maltose, and mannose caused inhibitory effects similar to that observed with glucose, but ribose and 2-deoxyglucose did not affect the rate of malolactic activity. The addition of fructose or citrate completely relieved the glucose-induced inhibition. Glucose was not catabolized by permeabilized cells, and inhibition of malolactic fermentation was not observed under these conditions. 31P NMR analysis of perchloric acid extracts of cells obtained during glucose-malate cometabolism showed high intracellular concentrations of glucose-6-phosphate, 6-phosphogluconate, and glycerol-3-phosphate. Glucose-6-phosphate, 6-phosphogluconate, and NAD(P)H inhibited the malolactic activity in permeabilized cells or cell extracts, whereas NADP+ had no inhibitory effect. The purified malolactic enzyme was strongly inhibited by NADH, whereas all the other above-mentioned metabolites exerted no inhibitory effect, showing that NADH was responsible for the inhibition of malolactic activity in vivo. The concentration of NADH required to inhibit the activity of the malolactic enzyme by 50% was ca. 25 microM. The data provide a coherent biochemical basis to understand the glucose-induced inhibition of malolactic fermentation in L. oenos.

Coffman, JA; Cooper, TG

doi: N/Apmid: 9287023

JA Coffman and TG Cooper Department of Microbiology and Immunology, University of Tennessee, Memphis 38163, USA. The expression of most nitrogen catabolic genes in Saccharomyces cerevisiae is regulated at the level of transcription in response to the quality of nitrogen source available. This regulation is accomplished through four GATA-family transcription factors: two positively acting factors capable of transcriptional activation (Gln3p and Gat1p) and two negatively acting factors capable of down-regulating Gln3p- and/or Gat1p-dependent transcription (Dal80p and Deh1p). Current understanding of nitrogen-responsive transcriptional regulation is the result of extensive analysis of genes required for the catabolism of small molecules, e.g., amino acids, allantoin, or ammonia. However, cells contain another, equally important source of nitrogen, intracellular protein, which undergoes rapid turnover during special circumstances such as entry into stationary phase, and during sporulation. Here we show that the expression of some (CPS1, PEP4, PRB1, and LAP4) but not all (PRC1) vacuolar protease genes is nitrogen catabolite repression sensitive and is regulated by the GATA-family proteins Gln3p, Gat1p, and Dal80p. These observations extend the global participation of GATA-family transcription factors to include not only well-studied genes associated with the catabolism of small nitrogenous compounds but also genes whose products are responsible for the turnover of intracellular macromolecules. They also point to the usefulness of considering control of the nitrogen-responsive GATA factors when studying the regulation of the protein turnover machinery.

Wang, X; Huang, J; Mukherjee, A; Cao, C; Lutkenhaus, J

doi: N/Apmid: 9287012

Analysis of the interaction of FtsZ with itself, GTP, and FtsA. X Wang , J Huang , A Mukherjee , C Cao and J Lutkenhaus Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City 66160, USA. ABSTRACT The interaction of FtsZ with itself, GTP, and FtsA was examined by analyzing the sensitivity of FtsZ to proteolysis and by using the yeast two-hybrid system. The N-terminal conserved domain consisting of 320 amino acids bound GTP, and a central region of FtsZ, encompassing slightly more than half of the protein, was cross-linked to GTP. Site-directed mutagenesis revealed that none of six highly conserved aspartic acid and asparagine residues were required for GTP binding. These results indicate that the specificity determinants for GTP binding are different than those for the GTPase superfamily. The N-terminal conserved domain of FtsZ contained a site for self-interaction that is conserved between FtsZ proteins from distantly related bacterial species. FtsZ320, which was truncated at the end of the conserved domain, was a potent inhibitor of division although it expressed normal GTPase activity and could polymerize. FtsZ was also found to interact directly with FtsA, and this interaction could also be observed between these proteins from distantly related bacterial species. CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? « Previous | Next Article » Table of Contents This Article J. Bacteriol. September 1997 vol. 179 no. 17 5551-5559 » Abstract PDF Services Email this article to a colleague Similar articles in ASM journals Alert me when this article is cited Alert me if a correction is posted Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of JB Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Load citing article information Citing articles via Web of Science Citing articles via Google Scholar Google Scholar Articles by Wang, X. Articles by Lutkenhaus, J. Search for related content PubMed PubMed citation Articles by Wang, X. Articles by Lutkenhaus, J. Related Content Load related web page information Social Bookmarking CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? current issue December 2011, volume 193, issue 24 Alert me to new issues of JB About JB Subscribers Authors Reviewers Advertisers Inquiries from the Press Permissions & Commercial Reprints ASM Journals Public Access Policy JB RSS Feeds 1752 N Street N.W. • Washington DC 20036 202.737.3600 • 202.942.9355 fax • [email protected] Print ISSN: 0021-9193 Online ISSN: 1098-5530 Copyright © 2011 by the American Society for Microbiology. For an alternate route to JB .asm.org, visit: http://intl- JB .asm.org | More Info»

Mojica, FJ; Cisneros, E; Ferrer, C; Rodriguez-Valera, F; Juez, G

doi: N/Apmid: 9287003

FJ Mojica, E Cisneros, C Ferrer, F Rodriguez-Valera and G Juez Departamento Genetica y Microbiologia, Universidad de Alicante, Campus de San Juan, Spain. Haloferax volcanii and Halomonas elongata have been selected as representatives of halophilic Archaea and Bacteria, respectively, to analyze the responses to various osmolarities at the protein synthesis level. We have identified a set of high-salt-related proteins (39, 24, 20, and 15.5 kDa in H. elongata; 70, 68, 48, and 16 kDa in H. volcanii) whose synthesis rates increased with increasing salinities. A different set of proteins (60, 42, 15, and 6 kDa for H. elongata; 63, 44, 34, 18, 17, and 6 kDa for H. volcanii), some unique for low salinities, was induced under low-salt conditions. For both organisms, and especially for the haloarchaeon, adaptation to low-salt conditions involved a stronger and more specific response than adaptation to high-salt conditions, indicating that unique mechanisms may have evolved for low- salinity adaptation. In the case of H. volcanii, proteins with a typical transient response to osmotic shock, induced by both hypo- and hyperosmotic conditions, probably corresponding to described heat shock proteins and showing the characteristics of general stress proteins, have also been identified. Cell recovery after a shift to low salinities was immediate in both organisms. In contrast, adaptation to higher salinities in both cases involved a lag period during which growth and general protein synthesis were halted, although the high- salt-related proteins were induced rapidly. In H. volcanii, this lag period corresponded exactly to the time needed for cells to accumulate adequate intracellular potassium concentrations, while extrusion of potassium after the down-shift was immediate. Thus, reaching osmotic balance must be the main limiting factor for recovery of cell functions after the variation in salinity.

Wang, X; Huang, J; Mukherjee, A; Cao, C; Lutkenhaus, J

doi: N/Apmid: 9287012

X Wang, J Huang, A Mukherjee, C Cao and J Lutkenhaus Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City 66160, USA. The interaction of FtsZ with itself, GTP, and FtsA was examined by analyzing the sensitivity of FtsZ to proteolysis and by using the yeast two-hybrid system. The N-terminal conserved domain consisting of 320 amino acids bound GTP, and a central region of FtsZ, encompassing slightly more than half of the protein, was cross-linked to GTP. Site- directed mutagenesis revealed that none of six highly conserved aspartic acid and asparagine residues were required for GTP binding. These results indicate that the specificity determinants for GTP binding are different than those for the GTPase superfamily. The N- terminal conserved domain of FtsZ contained a site for self-interaction that is conserved between FtsZ proteins from distantly related bacterial species. FtsZ320, which was truncated at the end of the conserved domain, was a potent inhibitor of division although it expressed normal GTPase activity and could polymerize. FtsZ was also found to interact directly with FtsA, and this interaction could also be observed between these proteins from distantly related bacterial species.

Garrigues, C; Loubiere, P; Lindley, ND; Cocaign-Bousquet, M

doi: N/Apmid: 9286977

C Garrigues, P Loubiere, ND Lindley and M Cocaign-Bousquet Centre National de la Recherche Scientifique-Unite Mixte de Recherche 5504, Laboratoire Associe Institut National de la Recherche Agronomique, Institut National des Sciences Appliquees, Toulouse, France. During batch growth of Lactococcus lactis subsp. lactis NCDO 2118 on various sugars, the shift from homolactic to mixed-acid metabolism was directly dependent on the sugar consumption rate. This orientation of pyruvate metabolism was related to the flux-controlling activity of glyceraldehyde-3-phosphate dehydrogenase under conditions of high glycolytic flux on glucose due to the NADH/NAD+ ratio. The flux limitation at the level of glyceraldehyde-3-phosphate dehydrogenase led to an increase in the pool concentrations of both glyceraldehyde-3- phosphate and dihydroxyacetone-phosphate and inhibition of pyruvate formate lyase activity. Under such conditions, metabolism was homolactic. Lactose and to a lesser extent galactose supported less rapid growth, with a diminished flux through glycolysis, and a lower NADH/NAD+ ratio. Under such conditions, the major pathway bottleneck was most probably at the level of sugar transport rather than glyceraldehyde-3-phosphate dehydrogenase. Consequently, the pool concentrations of phosphorylated glycolytic intermediates upstream of glyceraldehyde-3-phosphate dehydrogenase decreased. However, the intracellular concentration of fructose-1,6-bisphosphate remained sufficiently high to ensure full activation of lactate dehydrogenase and had no in vivo role in controlling pyruvate metabolism, contrary to the generally accepted opinion. Regulation of pyruvate formate lyase activity by triose phosphates was relaxed, and mixed-acid fermentation occurred (no significant production of lactate on lactose) due mostly to the strong inhibition of lactate dehydrogenase by the in vivo NADH/NAD+ ratio.

Amster-Choder, O; Wright, A

doi: N/Apmid: 9287026

O Amster-Choder and A Wright Department of Molecular Biology, Hadassah Medical School, The Hebrew University, Jerusalem, Israel. We have shown previously that the activity of BglG, the response regulator of the bgl system, as a transcriptional antiterminator is modulated by the sensor BglF, which reversibly phosphorylates BglG. We show here that the phosphoryl group on BglG is present as a phosphoramidate, based on the sensitivity of phosphorylated BglG to heat, hydroxylamine, and acidic but not basic conditions. By analyzing the products of base-hydrolyzed phosphorylated BglG by thin-layer chromatography, we show that the phosphorylation occurs on a histidine residue. This result supports the notion that the bgl system is a member of a new family of bacterial sensory systems.

Coffman, J A; Cooper, T G

doi: N/Apmid: 9287023

Nitrogen GATA factors participate in transcriptional regulation of vacuolar protease genes in Saccharomyces cerevisiae. J A Coffman and T G Cooper Department of Microbiology and Immunology, University of Tennessee, Memphis 38163, USA. ABSTRACT The expression of most nitrogen catabolic genes in Saccharomyces cerevisiae is regulated at the level of transcription in response to the quality of nitrogen source available. This regulation is accomplished through four GATA-family transcription factors: two positively acting factors capable of transcriptional activation (Gln3p and Gat1p) and two negatively acting factors capable of down-regulating Gln3p- and/or Gat1p-dependent transcription (Dal80p and Deh1p). Current understanding of nitrogen-responsive transcriptional regulation is the result of extensive analysis of genes required for the catabolism of small molecules, e.g., amino acids, allantoin, or ammonia. However, cells contain another, equally important source of nitrogen, intracellular protein, which undergoes rapid turnover during special circumstances such as entry into stationary phase, and during sporulation. Here we show that the expression of some (CPS1, PEP4, PRB1, and LAP4) but not all (PRC1) vacuolar protease genes is nitrogen catabolite repression sensitive and is regulated by the GATA-family proteins Gln3p, Gat1p, and Dal80p. These observations extend the global participation of GATA-family transcription factors to include not only well-studied genes associated with the catabolism of small nitrogenous compounds but also genes whose products are responsible for the turnover of intracellular macromolecules. They also point to the usefulness of considering control of the nitrogen-responsive GATA factors when studying the regulation of the protein turnover machinery. CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? « Previous | Next Article » Table of Contents This Article J. Bacteriol. September 1997 vol. 179 no. 17 5609-5613 » Abstract PDF Services Email this article to a colleague Similar articles in ASM journals Alert me when this article is cited Alert me if a correction is posted Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of JB Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Load citing article information Citing articles via Web of Science Citing articles via Google Scholar Google Scholar Articles by Coffman, J. A. Articles by Cooper, T. G. Search for related content PubMed PubMed citation Articles by Coffman, J. A. Articles by Cooper, T. G. Related Content Load related web page information Social Bookmarking CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? current issue December 2011, volume 193, issue 24 Alert me to new issues of JB About JB Subscribers Authors Reviewers Advertisers Inquiries from the Press Permissions & Commercial Reprints ASM Journals Public Access Policy JB RSS Feeds 1752 N Street N.W. • Washington DC 20036 202.737.3600 • 202.942.9355 fax • [email protected] Print ISSN: 0021-9193 Online ISSN: 1098-5530 Copyright © 2011 by the American Society for Microbiology. For an alternate route to JB .asm.org, visit: http://intl- JB .asm.org | More Info»