Identification of vegetative proteins for a two-dimensional protein index of Bacillus subtilis

Schmid, Roland; Bernhardt, Jörg; Antelmann, Haike; Völker, Andrea; Mach, Hiltraut; Völker, Uwe; Hecker, Michael

doi:10.1099/00221287-143-3-991

Cited by 66 publications

(42 citation statements)

References 21 publications

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“…Therefore, sequence selectivity in RNA binding is apparent for CSPs but may not be high. Schmid et al (1997) and Bernhardt et al (1997). Other spots identified in Graumann et al (1996).…”

Section: Loss Of Csps Leads To An Increase In Levels Of Remaining Cspmentioning

confidence: 97%

A family of cold shock proteins in Bacillus subtilis is essential for cellular growth and for efficient protein synthesis at optimal and low temperatures

Graumann

Wendrich

Weber

et al. 1997

Molecular Microbiology

233

266

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SummaryLike other bacteria, Bacillus subtilis possesses a family of homologous small acidic proteins (CspB, CspC and CspD, identity > 70%) that are strongly induced in response to cold shock. We show that deletion of cspC or cspD genes did not result in a detectable phenotype; in contrast, csp double mutants exhibited severe reduction in cellular growth at 15ЊC as well as at 37ЊC, including impairment of survival during the stationary phase. Two-dimensional gel analysis showed that protein synthesis was deregulated in csp double mutants and that the loss of one or two CSPs led to an increase in the synthesis of the remaining CSP(s) at 37ЊC and after cold shock, suggesting that CSPs down-regulate production of members from this protein family. A cspB/C/D triple mutant (64BCDbt) could only be generated in the presence of cspB in trans on a plasmid that was not lost, in spite of lack of antibiotic pressure, indicating that a minimum of one csp gene is essential for viability of B. subtilis. After cold shock, synthesis of CspB in 64BCDbt was drastically lower than in wild-type cells accompanied by cessation in growth and strong reduction in general protein synthesis. As CspB, CspC and CspD are shown to bind to RNA in a co-operative and interactive manner, CSPs are suggested to function as RNA chaperones facilitating the initiation of translation under optimal and low temperatures.

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“…Therefore, sequence selectivity in RNA binding is apparent for CSPs but may not be high. Schmid et al (1997) and Bernhardt et al (1997). Other spots identified in Graumann et al (1996).…”

Section: Loss Of Csps Leads To An Increase In Levels Of Remaining Cspmentioning

confidence: 97%

A family of cold shock proteins in Bacillus subtilis is essential for cellular growth and for efficient protein synthesis at optimal and low temperatures

Graumann

Wendrich

Weber

et al. 1997

Molecular Microbiology

233

266

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“…The resulting protein spot was blotted onto a polyvinylidene difluoride membrane (Immobilon P) using a buffer system consisting of 10 mM NaHCO 3 and 3 mM Na 2 CO 3 . After staining with Amido Black, the spot was excised and installed into the blot cartridge of a model 473A protein sequencer (Applied Biosystems), and its amino acid sequence was determined as described previously (11).…”

Section: Methodsmentioning

confidence: 99%

A Novel Insect V-ATPase Subunit M9.7 Is Glycosylated Extensively

Merzendorfer

Huss

Schmid

et al. 1999

Journal of Biological Chemistry

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Plasma membrane V-ATPase isolated from midgut and Malpighian tubules of the tobacco hornworm, Manduca sexta, contains a novel prominent 20-kDa polypeptide. Based on N-terminal protein sequencing, we cloned a corresponding cDNA. The deduced hydrophobic protein consisted of 88 amino acids with a molecular mass of only 9.7 kDa. Immunoblots of the recombinant 9.7-kDa polypeptide, using a monoclonal antibody to the 20-kDa polypeptide, confirmed that the correct cDNA had been cloned. The 20-kDa polypeptide is glycosylated, as deduced from lectin staining. Treatment with N-glycosidase A resulted in the appearance of two additional protein bands of 16 and 10 kDa which both were immunoreactive to the 20-kDa polypeptidespecific monoclonal antibody. Thus, extensive N-glycosylation of the novel V o subunit M9.7 accounts for half of its molecular mass observed in SDS-polyacrylamide gel electrophoresis. M9.7 exhibits some similarities to the yeast protein Vma21p which resides in the endoplasmic reticulum and is required for the assembly of the V o complex. However, as deduced from immunoblots as well as from activities of the V-ATPase and endoplasmic reticulum marker enzymes in different membrane preparations, M9.7 is, in contrast to the yeast polypeptide, a constitutive subunit of the mature plasma membrane V-ATPase of M. sexta. Hϩ V-ATPases are a class of ion transport proteins that couple ATP hydrolysis to the movement of protons across membranes. In endomembranes they function, in concert with chloride channels, as acidifiers of intracellular compartments, whereas in plasma membranes their roles are dependent on the cell type. V-ATPases consist of a peripheral V 1 complex, which is responsible for the hydrolysis of ATP, and a membranebound V o complex which is responsible for the translocation of protons. Although the subunit composition may depend upon the source of the enzyme, at least seven subunits of the V 1 complex, subunits A to G, appear to be universal V-ATPase components (1). By contrast, the subunit composition of the V o complex is less clear. There is no doubt that a 16 -17-kDa proteolipid, the proton "channel," is a major constituent of the V o complex. A membrane-associated subunit in the 40-kDa range and an ϳ100-kDa transmembrane subunit may be two additional essential V o components (1). Recently, a novel 9.2-kDa membrane sector-associated polypeptide was reported from bovine chromaffin granules (2). Its sequence and structure show some similarity to Vma21p, a yeast protein involved in the assembly of the V-ATPase; whether or not it is a constitutive V-ATPase subunit remains an open question.In the larval midgut epithelium of the model insect, Manduca sexta (Lepidoptera, Sphingidae), a plasma membrane V-ATPase is present in the apical membranes of goblet cells where it energizes the alkalinization of the gut lumen to a pH of more than 11 (3). For the V 1 complex, amino acid sequences of five insect subunits A, B, E, F, and G have been deduced from cloned cDNAs (4), and evidence for the existence of subuni...

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“…The accompanying paper (Schmid et al, 1997) presents a catalogue of known vegetative proteins from our 2-D gels. Using these known vegetative proteins as marker spots, a 2-D master gel was established and the protein synthesis pattern during exponential growth under optimal laboratory conditions is described in the accompanying paper (Schmid et al,1.997 Volker et al, 1995a) were cultivated in a synthetic medium described previously (Stiilke et al, 1993).…”

Section: Gspl6mentioning

confidence: 99%

Specific and general stress proteins in Bacillus subtilis � a two-dimensional protein electrophoresis study

et al. 1997

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A computer-aided analysis of high resolution two-dimensional polyacrylamide gels was used to investigate the changes in the protein synthesis profile in B. subtilis wild-type strains and sigB mutants in response to heat shock, salt and ethanol stress, and glucose or phosphate starvation. The data provided evidence that the induction of a t least 42 general stress proteins absolutely required the alternative sigma factor oB. However, a t least seven stress proteins, among them ClpC, ClpP, Sod, AhpC and AhpF, remained stressinducible in a sigB mutant. Such a detailed analysis also permitted the description of subgroups of general stress proteins which are subject to additional regulatory circuits, indicating a very thorough fine-tuning of this complex response. The relative synthesis rate of the general stress proteins constituted up to 40% of the total protein synthesis of stressed cells and thereby emphasizes the importance of the stress regulon. Besides the induction of these general or rather unspecific stress proteins, the induction of stress-specif ic proteins is shown and discussed.

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Identification of vegetative proteins for a two-dimensional protein index of Bacillus subtilis

Cited by 66 publications

References 21 publications

A family of cold shock proteins in Bacillus subtilis is essential for cellular growth and for efficient protein synthesis at optimal and low temperatures

A family of cold shock proteins in Bacillus subtilis is essential for cellular growth and for efficient protein synthesis at optimal and low temperatures

A Novel Insect V-ATPase Subunit M9.7 Is Glycosylated Extensively

Specific and general stress proteins in Bacillus subtilis � a two-dimensional protein electrophoresis study

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