scholarly journals A Region in Bacillus subtilisςH Required for Spo0A-Dependent Promoter Activity

1998 ◽  
Vol 180 (18) ◽  
pp. 4987-4990 ◽  
Author(s):  
Cindy M. Buckner ◽  
Charles P. Moran
Keyword(s):  
Amino Acids ◽  
Bacillus Subtilis ◽  
Amino Acid ◽  
Rna Polymerase ◽  
Sigma Factor ◽  
Promoter Activity ◽  
Amino Acid Substitutions ◽  
Mutant Form ◽  
Promoter Activation ◽  
Affect Expression

ABSTRACT Spo0A activates transcription in Bacillus subtilis from promoters that are used by two types of RNA polymerase, RNA polymerase containing the primary sigma factor, ςA, and RNA polymerase containing a secondary sigma factor, known as ςH. The region of ςA near positions 356 to 359 is required for Spo0A-dependent promoter activation, possibly because Spo0A interacts with this region of ςA at these promoters. To determine if the amino acids in the corresponding region of ςH are also important in Spo0A-dependent promoter activation, we examined the effects of single alanine substitutions at 10 positions in ςH (201 to 210). Two alanine substitutions in ςH, at glutamine 201 (Q201A) and at arginine 205 (R205A), significantly decreased activity from the Spo0A-dependent, ςH-dependent promoterspoIIA but did not affect expression from the ςH-dependent, Spo0A-independent promoterscitGp2 and spoVG. Therefore, promoter activation by Spo0A requires homologous regions in ςA and ςH. A mutant form of Spo0A, S231F, that suppresses the sporulation defect caused by several amino acid substitutions in ςA did not suppress the sporulation defects caused by the Q201A and R205A substitutions in ςH. This result and others indicate that different surfaces of Spo0A probably interact with ςA and ςH RNA polymerases.

scholarly journals α-Helix E of Spo0A Is Required for σA- but Not for σH-Dependent Promoter Activation in Bacillus subtilis

2004 ◽  
Vol 186 (4) ◽  
pp. 1078-1083 ◽  
Author(s):  
Amrita Kumar ◽  
James A. Brannigan ◽  
Charles P. Moran
Keyword(s):  
Amino Acids ◽  
Bacillus Subtilis ◽  
Amino Acid ◽  
Rna Polymerase ◽  
Sigma Factor ◽  
Dna Binding Protein ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Promoter Activation ◽  
Α Helix

ABSTRACT At the onset of endospore formation in Bacillus subtilis, the DNA binding protein Spo0A activates transcription from two types of promoters. The first type includes the spoIIG and spoIIE promoters, which are used by σA-RNA polymerase, whereas the second type includes the spoIIA promoter, which is used by RNA polymerase containing the secondary sigma factor σH. Previous genetic analyses have identified specific amino acids in α-helix E of Spo0A that are important for activation of Spo0A-dependent, σA-dependent promoters. However, these amino acids are not required for activation of the σH-dependent spoIIA promoter. We now report the effects of additional single-amino-acid substitutions and the effects of deletions in α-helix E. The effects of alanine substitutions revealed one new position (239) in Spo0A that appears to be specifically required for activation of the σA-dependent promoters. Based on the effects of a deletion mutation, we suggest that α-helix E in Spo0A is not directly involved in interaction with σH-RNA polymerase.

scholarly journals Surfaces of Spo0A and RNA Polymerase Sigma Factor A That Interact at the spoIIG Promoter in Bacillus subtilis

2004 ◽  
Vol 186 (1) ◽  
pp. 200-206 ◽  
Author(s):  
Amrita Kumar ◽  
Cindy Buckner Starke ◽  
Mark DeZalia ◽  
Charles P. Moran
Keyword(s):  
Bacillus Subtilis ◽  
Amino Acid ◽  
Rna Polymerase ◽  
Sigma Factor ◽  
Dna Binding Protein ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
Promoter Activation ◽  
Interacting Surfaces

ABSTRACT In Bacillus subtilis, the DNA binding protein Spo0A activates transcription from two classes of promoters, those used by RNA polymerase containing the primary sigma factor, σA (e.g., spoIIG), and those used by RNA polymerase containing the secondary sigma factor, σH (e.g., spoIIA). Several single amino acid substitutions in region 4 of σA define positions in σA that are specifically required for Spo0A-dependent promoter activation. Similarly, several single amino acid substitutions in Spo0A define positions in Spo0A that are required for σA-dependent promoter activation but not for other functions of Spo0A. It is unknown whether these amino acids in Spo0A interact directly with those in region 4 of σA or whether they interact with another subunit of RNA polymerase to effect promoter activation. Here we report the identification of a new amino acid in region 4 of σA, arginine at position 355 (R355), that is involved in Spo0A-dependent promoter activation. To further investigate the role of R355, we used the coordinates of Spo0A and sigma region 4, each in complex with DNA, to build a model for the interaction of σA and Spo0A at the spoIIG promoter. We tested the model by examining the effects of amino acid substitutions in the putative interacting surfaces of these molecules. As predicted by the model, we found genetic evidence for interaction of R355 of σA with glutamine at position 221 of Spo0A. These results appear to define the surfaces of Spo0A and σA that directly interact during activation of the spoIIG promoter.

scholarly journals A Region of ςK Involved in Promoter Activation by GerE in Bacillus subtilis

1999 ◽  
Vol 181 (14) ◽  
pp. 4365-4373 ◽  
Author(s):  
Kathryn H. Wade ◽  
Ghislain Schyns ◽  
Jason A. Opdyke ◽  
Charles P. Moran
Keyword(s):  
Bacillus Subtilis ◽  
Amino Acid ◽  
Rna Polymerase ◽  
Promoter Activity ◽  
Amino Acid Substitutions ◽  
Amino Acid Residues ◽  
Transcription In Vitro ◽  
A Site ◽  
Initial Binding

ABSTRACT During endospore formation in Bacillus subtilis, the DNA binding protein GerE stimulates transcription from several promoters that are used by RNA polymerase containing ςK. GerE binds to a site on one of these promoters, cotX, that overlaps its −35 region. We tested the model that GerE interacts with ςK at the cotX promoter by seeking amino acid substitutions in ςK that interfered with GerE-dependent activation of the cotX promoter but which did not affect utilization of the ςK-dependent, GerE-independent promoter gerE. We identified two amino acid substitutions in ςK, E216K and H225Y, that decrease cotXpromoter utilization but do not affect gerE promoter activity. Alanine substitutions at these positions had similar effects. We also examined the effects of the E216A and H225Y substitutions in ςK on transcription in vitro. We found that these substitutions specifically reduced utilization of the cotXpromoter. These and other results suggest that the amino acid residues at positions 216 and 225 are required for GerE-dependentcotX promoter activity, that the histidine at position 225 of ςK may interact with GerE at the cotXpromoter, and that this interaction may facilitate the initial binding of ςK RNA polymerase to the cotX promoter. We also found that the alanine substitutions at positions 216 and 225 of ςK had no effect on utilization of the GerE-dependent promoter cotD, which contains GerE binding sites that do not overlap with its −35 region.

scholarly journals ςK Can Negatively RegulatesigE Expression by Two Different Mechanisms during Sporulation of Bacillus subtilis

1999 ◽  
Vol 181 (13) ◽  
pp. 4081-4088 ◽  
Author(s):  
Bin Zhang ◽  
Paolo Struffi ◽  
Lee Kroos
Keyword(s):  
Bacillus Subtilis ◽  
Rna Polymerase ◽  
Sigma Factor ◽  
Mother Cell ◽  
Transcriptional Activity ◽  
The Other ◽  
Mutant Form ◽  
Gene Encoding ◽  
Negative Effect ◽  
Cell Gene Expression

ABSTRACT Temporal and spatial gene regulation during Bacillus subtilis sporulation involves the activation and inactivation of multiple sigma subunits of RNA polymerase in a cascade. In the mother cell compartment of sporulating cells, expression of thesigE gene, encoding the earlier-acting sigma factor, ςE, is negatively regulated by the later-acting sigma factor, ςK. Here, it is shown that the negative feedback loop does not require SinR, an inhibitor of sigEtranscription. Production of ςK about 1 h earlier than normal does affect Spo0A, which when phosphorylated is an activator of sigE transcription. A mutation in thespo0A gene, which bypasses the phosphorelay leading to the phosphorylation of Spo0A, diminished the negative effect of early ςK production on sigE expression early in sporulation. Also, early production of ςK reduced expression of other Spo0A-dependent genes but not expression of the Spo0A-independent ald gene. In contrast, bothsigE and ald were overexpressed late in development of cells that fail to make ςK. Theald promoter, like the sigE promoter, is believed to be recognized by ςA RNA polymerase, suggesting that ςK may inhibit ςA activity late in sporulation. To exert this negative effect, ςKmust be transcriptionally active. A mutant form of ςKthat associates with core RNA polymerase, but does not direct transcription of a ςK-dependent gene, failed to negatively regulate expression of sigE or aldlate in development. On the other hand, the negative effect of early ςK production on sigE expression early in sporulation did not require transcriptional activity of ςK RNA polymerase. These results demonstrate that ςK can negatively regulate sigE expression by two different mechanisms, one observed when ςK is produced earlier than normal, which does not require ςKto be transcriptionally active and affects Spo0A, and the other observed when ςK is produced at the normal time, which requires ςK RNA polymerase transcriptional activity. The latter mechanism facilitates the switch from ςE to ςK in the cascade controlling mother cell gene expression.

scholarly journals Identification of separate domains in the adenovirus E1A gene for immortalization activity and the activation of virus early genes.

1986 ◽  
Vol 6 (10) ◽  
pp. 3470-3480 ◽  
Author(s):  
E Moran ◽  
B Zerler ◽  
T M Harrison ◽  
M B Mathews
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Point Mutations ◽  
Apparent Molecular Weight ◽  
Amino Acid Position ◽  
Cysteine Residue ◽  
Transformation Function ◽  
Single Amino Acid ◽  
Amino Acid Substitutions ◽  
E1a Gene

The transformation and early adenovirus gene transactivation functions of the E1A region were analyzed with deletion and point mutations. Deletion of amino acids from position 86 through 120 had little effect on the lytic or transforming functions of the E1A products, while deletion of amino acids from position 121 through 150 significantly impaired both functions. The sensitivity of the transformation function to alterations in the region from amino acid position 121 to 150 was further indicated by the impairment of transforming activity resulting from single amino acid substitutions at positions 124 and 135. Interestingly, conversion of a cysteine residue at position 124 to glycine severely impaired the transformation function without affecting the early adenovirus gene activating functions. Single amino acid substitutions in a different region of the E1A gene had the converse effect. All the mutants produced polypeptides of sufficient stability to be detected by Western immunoblot analysis. The single amino acid substitutions at positions 124 and 135, although impairing the transformation functions, did not detectably alter the formation of the higher-apparent-molecular-weight forms of the E1A products.

PREDICTION OF PATHOGENIC EFFECT FOR AMINO ACID SUBSTITUTIONS IN BRCA1 AND BRCA2 PROTEINS USING MNA DESCRIPTORS AND NAIVE BAYES CLASSIFIER

2020 ◽  
pp. 250-252
Author(s):  
D. Filimonov ◽  
A. Lagunin
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Sequence ◽  
Amino Acid Substitutions ◽  
Classification Models ◽  
Bayes Classifier ◽  
Brca1 And Brca2 ◽  
Chemical Structures ◽  
Pathogenic Effect ◽  
Pathogenic Effects

It is advisable to use data peptide's chemical structures with amino acids (AMA) substitution and the corresponding sections of the protein sequence without mutation to construct classification models predicting the pathogenic effects AMA substitutions based on MNA descriptors.

scholarly journals Genome-Wide Analysis of the Stationary-Phase Sigma Factor (Sigma-H) Regulon of Bacillus subtilis

2002 ◽  
Vol 184 (17) ◽  
pp. 4881-4890 ◽  
Author(s):  
Robert A. Britton ◽  
Patrick Eichenberger ◽  
Jose Eduardo Gonzalez-Pastor ◽  
Paul Fawcett ◽  
Rita Monson ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Rna Polymerase ◽  
Stationary Phase ◽  
Sigma Factor ◽  
Transcriptional Profiling ◽  
Open Reading Frames ◽  
Nutrient Sources ◽  
Genome Wide ◽  
A Genome ◽  
Sigma H

ABSTRACT Sigma-H is an alternative RNA polymerase sigma factor that directs the transcription of many genes that function at the transition from exponential growth to stationary phase in Bacillus subtilis. Twenty-three promoters, which drive transcription of 33 genes, are known to be recognized by sigma-H-containing RNA polymerase. To identify additional genes under the control of sigma-H on a genome-wide basis, we carried out transcriptional profiling experiments using a DNA microarray containing >99% of the annotated B. subtilis open reading frames. In addition, we used a bioinformatics-based approach aimed at the identification of promoters recognized by RNA polymerase containing sigma-H. This combination of approaches was successful in confirming most of the previously described sigma-H-controlled genes. In addition, we identified 26 putative promoters that drive expression of 54 genes not previously known to be under the direct control of sigma-H. Based on the known or inferred function of most of these genes, we conclude that, in addition to its previously known roles in sporulation and competence, sigma-H controls genes involved in many physiological processes associated with the transition to stationary phase, including cytochrome biogenesis, generation of potential nutrient sources, transport, and cell wall metabolism.

scholarly journals Generation of Recombinant Rotavirus with an Antigenic Mosaic of Cross-Reactive Neutralization Epitopes on VP4

2008 ◽  
Vol 82 (13) ◽  
pp. 6753-6757 ◽  
Author(s):  
Satoshi Komoto ◽  
Masanori Kugita ◽  
Jun Sasaki ◽  
Koki Taniguchi
Keyword(s):  
Amino Acids ◽  
Monoclonal Antibodies ◽  
Amino Acid ◽  
Reverse Genetics ◽  
Amino Acid Substitutions ◽  
Neutralization Epitope ◽  
First Report ◽  
P Type

ABSTRACT Recombinant rotavirus (RV) with cDNA-derived chimeric VP4 was generated using recently developed reverse genetics for RV. The rescued virus, KU//rVP4(SA11)-II(DS-1), contains SA11 (simian RV strain, G3P[2])-based VP4, in which a cross-reactive neutralization epitope (amino acids 381 to 401) on VP5* is replaced by the corresponding sequence of a different P-type DS-1 (human RV strain, G2P[4]). Serological analyses with a panel of anti-VP4- and -VP7-neutralizing monoclonal antibodies revealed that the rescued virus carries a novel antigenic mosaic of cross-reactive neutralization epitopes on its VP4 surface. This is the first report of the generation of a recombinant RV with artificial amino acid substitutions.

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