Difference between revisions of "Sandbox"

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−
* '''Description:''' Glyceraldehyde 3-phosphate dehydrogenase, NAD-dependent, glycolytic enzyme <br/><br/>
+
* '''Description:''' phosphoglycerate kinase, glycolytic/ gluconeogenic enzyme<br/><br/>
  
 
{| align="right" border="1" cellpadding="2"  
 
{| align="right" border="1" cellpadding="2"  
 
|-
 
|-
 
|style="background:#ABCDEF;" align="center"|'''Gene name'''
 
|style="background:#ABCDEF;" align="center"|'''Gene name'''
−
|''gapA''
+
|''pgk''
 
|-
 
|-
 
|style="background:#ABCDEF;" align="center"| '''Synonyms''' || '' ''
 
|style="background:#ABCDEF;" align="center"| '''Synonyms''' || '' ''
 
|-
 
|-
−
|style="background:#ABCDEF;" align="center"| '''Essential''' || Yes [http://www.ncbi.nlm.nih.gov/sites/entrez/17114254 (PubMed)]
+
|style="background:#ABCDEF;" align="center"| '''Essential''' || yes
 
|-
 
|-
−
|style="background:#ABCDEF;" align="center"| '''Product''' || glyceraldehyde 3-phosphate dehydrogenase
+
|style="background:#ABCDEF;" align="center"| '''Product''' || phosphoglycerate kinase
 
|-
 
|-
−
|style="background:#ABCDEF;" align="center"|'''Function''' || catabolic enzyme in glycolysis
+
|style="background:#ABCDEF;" align="center"|'''Function''' || enzyme in glycolysis/ gluconeogenesis
 
|-
 
|-
−
|style="background:#ABCDEF;" align="center"| '''MW, pI''' || 35.7 kDa, 5.03
+
|style="background:#ABCDEF;" align="center"| '''MW, pI''' || 42,0 kDa, 4.77
 
|-
 
|-
−
|style="background:#ABCDEF;" align="center"| '''Gene length, protein length''' || 1005 bp, 335 amino acids
+
|style="background:#ABCDEF;" align="center"| '''Gene length, protein length''' || 1182 bp, 394 amino acids
 
|-
 
|-
−
|style="background:#ABCDEF;" align="center"|'''Immediate neighbours''' || ''[[cggR]]'', ''[[pgk]]''
+
|style="background:#ABCDEF;" align="center"|'''Immediate neighbours''' || ''[[gapA]]'', ''[[tpi]]''
 
|-
 
|-
 
|colspan="2" style="background:#FAF8CC;" align="center"|'''Hier soll was neues rein'''
 
|colspan="2" style="background:#FAF8CC;" align="center"|'''Hier soll was neues rein'''
 
|-
 
|-
−
|colspan="2" | '''Genetic context''' <br/> [[Image:gapA_context.gif]]
+
|colspan="2" | '''Genetic context''' <br/> [[Image:pgk_context.gif]]
 
  <div align="right"> <small>This image was kindly provided by [http://genolist.pasteur.fr/SubtiList/ SubtiList]</small></div>
 
  <div align="right"> <small>This image was kindly provided by [http://genolist.pasteur.fr/SubtiList/ SubtiList]</small></div>
 
|-
 
|-
Line 31: Line 31:
 
<br/><br/>
 
<br/><br/>
  
−
 
 
=The gene=
 
=The gene=
  
 
=== Basic information ===
 
=== Basic information ===
  
−
* '''Coordinates:''' 3480732 - 3481736
+
* '''Coordinates:''' 3479231 - 3480412
  
 
===Phenotypes of a mutant ===
 
===Phenotypes of a mutant ===
  
−
essential  [http://www.ncbi.nlm.nih.gov/pubmed/17114254 PubMed]
+
essential  [http://www.ncbi.nlm.nih.gov/pubmed/12682299 PubMed]
  
 
=== Database entries ===
 
=== Database entries ===
Line 46: Line 45:
 
* '''DBTBS entry:''' [http://dbtbs.hgc.jp/COG/prom/cggR-gapA-pgk-tpiA-pgm-eno.html]
 
* '''DBTBS entry:''' [http://dbtbs.hgc.jp/COG/prom/cggR-gapA-pgk-tpiA-pgm-eno.html]
  
−
* '''SubtiList entry:'''[http://genolist.pasteur.fr/SubtiList/genome.cgi?gene_detail+BG10827]
+
* '''SubtiList entry:''' [http://genolist.pasteur.fr/SubtiList/genome.cgi?gene_detail+BG11062]
  
 
=== Additional information===
 
=== Additional information===
Line 54: Line 53:
 
=== Basic information/ Evolution ===
 
=== Basic information/ Evolution ===
  
−
* '''Catalyzed reaction/ biological activity:''' glyceraldehyde-3-phosphate dehydrogenase, (NADH-dependent). Catalyzes the reaction from glyceraldehyde-3-phosphate to 1.3-bi-phosphoglycerate. This reaction is part of the glycolysis.
+
* '''Catalyzed reaction/ biological activity:''' ATP + 3-phospho-D-glycerate = ADP + 1,3-bisphosphoglycerate
  
−
* '''Protein family:'''
+
* '''Protein family:''' phosphoglycerate kinase family
  
−
* '''Paralogous protein(s):''' [[GapB]]
+
* '''Paralogous protein(s):'''
  
 
=== Extended information on the protein ===
 
=== Extended information on the protein ===
  
−
* '''Kinetic information:''' K(M) for NAD: 5.7 mM, K(cat) for NAD: 70/sec (determined for GapA from ''Geobacillus stearothermophilus'') [http://www.ncbi.nlm.nih.gov/sites/entrez/10799476 PubMed]
+
* '''Kinetic information:'''
  
 
* '''Domains:'''  
 
* '''Domains:'''  
 +
** nucleotide binding domain (ATP) (350–353)
 +
** 2x substrate binding domain (21–23), (59–62)
  
−
* '''Modification:''' phosphorylation on (Ser-148 OR Ser-151 OR Thr-153 OR Thr-154) [http://www.ncbi.nlm.nih.gov/sites/entrez/17218307 PubMed], [http://www.ncbi.nlm.nih.gov/pubmed/17726680 PubMed]
+
* '''Modification:''' phosphorylation on Ser-183 AND Thr-299 [http://www.ncbi.nlm.nih.gov/sites/entrez/17218307 PubMed], [http://www.ncbi.nlm.nih.gov/pubmed/17726680 PubMed]
  
 
* '''Cofactor(s):'''
 
* '''Cofactor(s):'''
Line 73: Line 74:
  
 
* '''Interactions:'''  
 
* '''Interactions:'''  
−
** GapA-[[PtsH]]: [[PtsH|HPr(Ser-46-P)]] binds GapA resulting in a slight inhibition of enzymatic activity.[http://www.ncbi.nlm.nih.gov/sites/entrez/17142398 PubMed]
 
−
** GapA-[[Crh]]: [[Crh|Crh(Ser-46-P)]] binds GapA resulting in a slight inhibition of enzymatic activity.[http://www.ncbi.nlm.nih.gov/sites/entrez/17142398 PubMed]
 
  
−
* '''Localization:''' Cytoplasm (Homogeneous) [http://www.ncbi.nlm.nih.gov/sites/entrez/16479537 PubMed] [http://www.ncbi.nlm.nih.gov/sites/entrez/14600241 PubMed], loosely membrane associated [http://www.ncbi.nlm.nih.gov/pubmed/18763711 PubMed]
+
* '''Localization:''' Cytoplasm (Homogeneous) [http://www.ncbi.nlm.nih.gov/sites/entrez/16479537 PubMed]
  
 
=== Database entries ===
 
=== Database entries ===
  
−
* '''Structure:'''  
+
* '''Structure:''' ''Geobacillus stearothermophilus'' [http://www.ncbi.nlm.nih.gov/Structure/mmdb/mmdbsrv.cgi?Dopt=s&uid=57113 NCBI]
−
** [http://www.rcsb.org/pdb/cgi/explore.cgi?pdbId=3CMC 3CMC] (from ''Geobacillus stearothermophilus'')
 
−
** [http://www.rcsb.org/pdb/cgi/explore.cgi?pdbId=1NQO 1NQO] (from ''Geobacillus stearothermophilus'', mutant with cys 149 replaced by ser, complex with NAD+ und D-Glyceraldehyde-3-Phosphate)
 
  
−
* '''Swiss prot entry:''' [http://www.expasy.ch/cgi-bin/sprot-search-ac?P09124 P09124]
+
* '''Swiss prot entry:''' [http://www.expasy.ch/cgi-bin/sprot-search-ac?P40924]
  
−
* '''KEGG entry:''' [http://www.genome.jp/dbget-bin/www_bget?bsu+BSU33940]
+
* '''KEGG entry:''' [http://www.genome.jp/dbget-bin/www_bget?bsu+BSU33930]
  
−
* '''E.C. number:''' [http://www.expasy.ch/cgi-bin/get-enzyme-entry?1.2.1.12 1.2.1.12]
+
* '''E.C. number:''' [http://www.expasy.org/enzyme/2.7.2.3 2.7.2.3]
  
 
=== Additional information===
 
=== Additional information===
  
−
GAP dehydrogenases from different sources (incl. ''Geobacillus stearothermophilus'') were shown to cleave RNA ([http://www.ncbi.nlm.nih.gov/sites/entrez/12359717 PubMed]). Moreover, mutations in ''gapA'' from ''B. subtilis'' can suppress mutations in genes involved in DNA replication ([http://www.ncbi.nlm.nih.gov/sites/entrez/17505547 PubMed]).
 
  
 
=Expression and regulation=
 
=Expression and regulation=
  
 
* '''Operon:'''  
 
* '''Operon:'''  
−
** ''[[cggR]]-[[gapA]]-[[pgk]]-[[tpi]]-[[pgm]]-[[eno]]''
+
** ''[[cggR]]-[[gapA]]-[[pgk]]-[[tpiA]]-[[pgm]]-[[eno]]''
−
** ''[[cggR]]-[[gapA]]''
+
** ''[[pgk]]-[[tpiA]]-[[pgm]]-[[eno]]''
−
 
 
−
The primary mRNAs of the operon are highly unstable. The primary mRNA is subject to processing at the very end of the ''[[cggR]]'' open reading frame. This results in stable mature ''[[gapA]]'' and ''[[gapA]]-[[pgk]]-[[tpiA]]-[[pgm]]-[[eno]]'' mRNAs. The processing event requires the [[Rny]] protein.
 
−
 
 
−
* '''Sigma factor:''' [[SigA]]
 
  
−
* '''Regulation:''' expression activated by glucose (10 fold) [http://www.ncbi.nlm.nih.gov/pubmed/12850135 PubMed],  [[CggR]] represses the operon in the absence of glycolytic sugars [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Abstract&list_uids=+12622823 PubMed]
+
* '''Sigma factor:''' [[SigA]]
  
−
* '''Regulatory mechanism:''' repression
+
* '''Regulation:''' expression activated by glucose (2.8 fold) [http://www.ncbi.nlm.nih.gov/pubmed/12850135 PubMed] 
 +
**''[[cggR]]'': induced by sugar [[CggR]] [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Abstract&list_uids=+11489127 PubMed]
 +
**''[[pgk]]'': constitutive [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Abstract&list_uids=+11489127 PubMed]
  
−
* '''Database entries:''' [http://dbtbs.hgc.jp/COG/prom/cggR-gapA-pgk-tpiA-pgm-eno.html DBTBS]
+
* '''Regulatory mechanism:''' transcription repression by [[CggR]] [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Abstract&list_uids=+11489127 PubMed]
  
−
* '''Additional information:''' GapA is one of the most abundant proteins in the cell. In the presence of glucose, there are about 25,000 GapA molecules per cell ([http://www.ncbi.nlm.nih.gov/sites/entrez/12634343 PubMed]).
+
* '''Additional information:'''
  
 
=Biological materials =
 
=Biological materials =
  
−
* '''Mutant:''' essential
+
* '''Mutant:'''
  
−
* '''Expression vector:''' pGP90 (N-terminal Strep-tag, purification from ''B. subtilis'', in [[pGP380]]), pGP704 (N-terminal His-tag, in [[pWH844]]) (available in [[Stülke]] lab)
+
* '''Expression vector:''' pGP1102 (N-terminal His-tag, in [[pWH844]]), pGP95 (N-terminal Strep-tag, in [[pGP172]]), pGP91 (N-terminal Strep-tag, for SPINE, expression in ''B. subtilis'', in [[pGP380]]), available in [[Stülke]] lab
 
 
−
* '''lacZ fusion:''' pGP506 (in [[pAC7]]), pGP512 (in [[pAC6]]) (available in [[Stülke]] lab)
+
* '''lacZ fusion:''' pGP514 (in [[pAC6]]), a series of promoter deletion variants is also available in [[pAC6]], available in [[Stülke]] lab
  
 
* '''GFP fusion:'''
 
* '''GFP fusion:'''
Line 124: Line 118:
 
* '''two-hybrid system:''' ''B. pertussis'' adenylate cyclase-based bacterial two hybrid system ([[BACTH]]), available in [[Stülke]] lab
 
* '''two-hybrid system:''' ''B. pertussis'' adenylate cyclase-based bacterial two hybrid system ([[BACTH]]), available in [[Stülke]] lab
  
−
* '''Antibody:''' available in [[Stülke]] lab
+
* '''Antibody:'''
  
 
=Labs working on this gene/protein=
 
=Labs working on this gene/protein=
−
 
−
[[Stephane Aymerich |Stephane Aymerich]], Microbiology and Molecular Genetics, INRA Paris-Grignon, France
 
−
 
−
[[Stülke|Jörg Stülke]], University of Göttingen, Germany
 
−
[http://wwwuser.gwdg.de/~genmibio/stuelke.html homepage]
 
  
 
=Your additional remarks=
 
=Your additional remarks=
Line 140: Line 129:
 
# Eymann et al. (2007) Dynamics of protein phosphorylation on Ser/Thr/Tyr in ''Bacillus subtilis''. ''Proteomics'' '''7:''' 3509-3526. [http://www.ncbi.nlm.nih.gov/pubmed/17726680 PubMed]
 
# Eymann et al. (2007) Dynamics of protein phosphorylation on Ser/Thr/Tyr in ''Bacillus subtilis''. ''Proteomics'' '''7:''' 3509-3526. [http://www.ncbi.nlm.nih.gov/pubmed/17726680 PubMed]
 
# Meile et al. (2006) Systematic localisation of proteins fused to the green fluorescent protein in ''Bacillus subtilis'': identification of new proteins at the DNA replication factory ''Proteomics'' '''6:''' 2135-2146. [http://www.ncbi.nlm.nih.gov/sites/entrez/16479537 PubMed]
 
# Meile et al. (2006) Systematic localisation of proteins fused to the green fluorescent protein in ''Bacillus subtilis'': identification of new proteins at the DNA replication factory ''Proteomics'' '''6:''' 2135-2146. [http://www.ncbi.nlm.nih.gov/sites/entrez/16479537 PubMed]
−
# Blencke, H.-M., Homuth, G., Ludwig, H., Mäder, U., Hecker, M. & Stülke, J. (2003) Transcriptional profiling of gene expression in response to glucose in Bacillus subtilis: regulation of the central metabolic pathways. Metab. Engn. 5: 133-149. [http://www.ncbi.nlm.nih.gov/sites/entrez/12850135 PubMed]
+
# Ludwig, H., Homuth, G., Schmalisch, M., Dyka, F. M., Hecker, M., and Stülke, J. (2001) Transcription of glycolytic genes and operons in ''Bacillus subtilis'': evidence for the presence of multiple levels of control of the ''gapA'' operon. Mol Microbiol 41, 409-422.[http://www.ncbi.nlm.nih.gov/sites/entrez/11489127 PubMed]
−
# Commichau, F. M., Rothe, F. M., Herzberg, C., Wagner, E., Hellwig, D., Lehnik-Habrink, M., Hammer, E., Völker, U. & Stülke, J. Novel activities of glycolytic enzymes in Bacillus subtilis: Interactions with essential proteins involved in mRNA processing. subm.
 
−
# Doan, T., and S. Aymerich. 2003. Regulation of the central glycolytic pathways in Bacillus subtilis: binding of the repressor CggR to its single DNA target sequence is modulated by fructose-1,6-bisphosphate. Mol. Microbiol. 47: 1709-1721. [http://www.ncbi.nlm.nih.gov/sites/entrez/12622823 PubMed]
 
−
# Evguenieva-Hackenberg, E., Schiltz, E., and Klug, G. (2002) Dehydrogenases from all three domains of life cleave RNA. J Biol Chem 277, 46145-46150. [http://www.ncbi.nlm.nih.gov/sites/entrez/12359717 PubMed]
 
−
# Fillinger, S., Boschi-Muller, S., Azza, S., Dervyn, E., Branlant, G., and Aymerich, S. (2000) Two glyceraldehyde-3-phosphate dehydrogenases with opposite physiological roles in a nonphotosynthetic bacterium. J Biol Chem 275, 14031-14037. [http://www.ncbi.nlm.nih.gov/sites/entrez/10799476 PubMed]
 
 
# Jannière, L., Canceill, D., Suski, C., Kanga, S., Dalmais, B., Lestini, R., Monnier, A. F., Chapuis, J., Bolotin, A., Titok, M., Le Chatelier, E., and Ehrlich, S. D. (2007) Genetic evidence for a link between glycolysis and DNA replication. PLoS ONE 2, e447. [http://www.ncbi.nlm.nih.gov/sites/entrez/17505547 PubMed]
 
# Jannière, L., Canceill, D., Suski, C., Kanga, S., Dalmais, B., Lestini, R., Monnier, A. F., Chapuis, J., Bolotin, A., Titok, M., Le Chatelier, E., and Ehrlich, S. D. (2007) Genetic evidence for a link between glycolysis and DNA replication. PLoS ONE 2, e447. [http://www.ncbi.nlm.nih.gov/sites/entrez/17505547 PubMed]
−
# Ludwig, H., Homuth, G., Schmalisch, M., Dyka, F. M., Hecker, M., and Stülke, J. (2001) Transcription of glycolytic genes and operons in ''Bacillus subtilis'': evidence for the presence of multiple levels of control of the ''gapA'' operon. Mol Microbiol 41, 409-422.[http://www.ncbi.nlm.nih.gov/sites/entrez/11489127 PubMed]
 
−
# Ludwig, H., Rebhan, N., Blencke, H.-M., Merzbacher, M. & Stülke, J. (2002). Control of the glycolytic ''gapA'' operon by the catabolite control protein A in ''Bacillus subtilis'': a novel mechanism of CcpA-mediated regulation. Mol Microbiol 45, 543-553.[http://www.ncbi.nlm.nih.gov/sites/entrez/12123463 PubMed]
 
 
# Macek et al. (2007) The serine/ threonine/ tyrosine phosphoproteome of the model  bacterium ''Bacillus subtilis''. Mol. Cell. Proteomics 6: 697-707  [http://www.ncbi.nlm.nih.gov/pubmed/17218307 PubMed]
 
# Macek et al. (2007) The serine/ threonine/ tyrosine phosphoproteome of the model  bacterium ''Bacillus subtilis''. Mol. Cell. Proteomics 6: 697-707  [http://www.ncbi.nlm.nih.gov/pubmed/17218307 PubMed]
−
# Meinken, C., Blencke, H. M., Ludwig, H., and Stülke, J. (2003) Expression of the glycolytic ''gapA'' operon in ''Bacillus subtilis'': differential synthesis of proteins encoded by the operon. Microbiology 149, 751-761. [http://www.ncbi.nlm.nih.gov/sites/entrez/12634343 PubMed]
 
−
# Pompeo ''et al.'' (2007) Interaction of GapA with HPr and its homologue, Crh: Novel levels of regulation of a key step of glycolysis in ''Bacillus subtilis''? J Bacteriol 189, 1154-1157.[http://www.ncbi.nlm.nih.gov/sites/entrez/17142398 PubMed]
 
−
# Thomaides, H. B., Davison, E. J., Burston, L., Johnson, H., Brown, D. R., Hunt, A. C., Errington, J., and Czaplewski, L. (2007) Essential bacterial functions encoded by gene pairs. J Bacteriol 189, 591-602. [http://www.ncbi.nlm.nih.gov/sites/entrez/17114254 PubMed]
 
−
# Tobisch, S., Zühlke, D., Bernhardt, J., Stülke, J., and Hecker, M. (1999) Role of CcpA in regulation of the central pathways of carbon catabolism in ''Bacillus subtilis''. J Bacteriol 181, 6996-7004.[http://www.ncbi.nlm.nih.gov/sites/entrez/10559165 PubMed]
 

Revision as of 17:44, 15 April 2009

  • Description: phosphoglycerate kinase, glycolytic/ gluconeogenic enzyme

Gene name pgk
Synonyms
Essential yes
Product phosphoglycerate kinase
Function enzyme in glycolysis/ gluconeogenesis
MW, pI 42,0 kDa, 4.77
Gene length, protein length 1182 bp, 394 amino acids
Immediate neighbours gapA, tpi
Hier soll was neues rein
Genetic context
Pgk context.gif
This image was kindly provided by SubtiList



The gene

Basic information

  • Coordinates: 3479231 - 3480412

Phenotypes of a mutant

essential PubMed

Database entries

  • DBTBS entry: [1]
  • SubtiList entry: [2]

Additional information

The protein

Basic information/ Evolution

  • Catalyzed reaction/ biological activity: ATP + 3-phospho-D-glycerate = ADP + 1,3-bisphosphoglycerate
  • Protein family: phosphoglycerate kinase family
  • Paralogous protein(s):

Extended information on the protein

  • Kinetic information:
  • Domains:
    • nucleotide binding domain (ATP) (350–353)
    • 2x substrate binding domain (21–23), (59–62)
  • Modification: phosphorylation on Ser-183 AND Thr-299 PubMed, PubMed
  • Cofactor(s):
  • Effectors of protein activity:
  • Interactions:
  • Localization: Cytoplasm (Homogeneous) PubMed

Database entries

  • Structure: Geobacillus stearothermophilus NCBI
  • Swiss prot entry: [3]
  • KEGG entry: [4]

Additional information

Expression and regulation

  • Regulatory mechanism: transcription repression by CggR PubMed
  • Additional information:

Biological materials

  • Mutant:
  • Expression vector: pGP1102 (N-terminal His-tag, in pWH844), pGP95 (N-terminal Strep-tag, in pGP172), pGP91 (N-terminal Strep-tag, for SPINE, expression in B. subtilis, in pGP380), available in Stülke lab
  • lacZ fusion: pGP514 (in pAC6), a series of promoter deletion variants is also available in pAC6, available in Stülke lab
  • GFP fusion:
  • two-hybrid system: B. pertussis adenylate cyclase-based bacterial two hybrid system (BACTH), available in Stülke lab
  • Antibody:

Labs working on this gene/protein

Your additional remarks

References

  1. Blencke et al. (2003) Transcriptional profiling of gene expression in response to glucose in Bacillus subtilis: regulation of the central metabolic pathways. Metab Eng. 5: 133-149 PubMed
  2. Eymann et al. (2007) Dynamics of protein phosphorylation on Ser/Thr/Tyr in Bacillus subtilis. Proteomics 7: 3509-3526. PubMed
  3. Meile et al. (2006) Systematic localisation of proteins fused to the green fluorescent protein in Bacillus subtilis: identification of new proteins at the DNA replication factory Proteomics 6: 2135-2146. PubMed
  4. Ludwig, H., Homuth, G., Schmalisch, M., Dyka, F. M., Hecker, M., and Stülke, J. (2001) Transcription of glycolytic genes and operons in Bacillus subtilis: evidence for the presence of multiple levels of control of the gapA operon. Mol Microbiol 41, 409-422.PubMed
  5. Jannière, L., Canceill, D., Suski, C., Kanga, S., Dalmais, B., Lestini, R., Monnier, A. F., Chapuis, J., Bolotin, A., Titok, M., Le Chatelier, E., and Ehrlich, S. D. (2007) Genetic evidence for a link between glycolysis and DNA replication. PLoS ONE 2, e447. PubMed
  6. Macek et al. (2007) The serine/ threonine/ tyrosine phosphoproteome of the model bacterium Bacillus subtilis. Mol. Cell. Proteomics 6: 697-707 PubMed
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