AlgA (PA3551) is the bifunctional phosphomannose isomerase / GDP-mannose pyrophosphorylase (PMI-GMP) of the alginate biosynthetic pathway, encoded as the last gene of the alginate operon. Its C-terminal type II PMI cupin domain converts fructose 6-phosphate to mannose 6-phosphate, which the phosphomannomutase AlgC converts to mannose 1-phosphate; its N-terminal nucleotidyltransferase domain then condenses mannose 1-phosphate with GTP to give GDP-mannose. GDP-mannose is oxidised by AlgD to GDP-mannuronic acid, the donor for alginate polymerisation. The purified enzyme is a monomer of about 54 kDa; the PMI reaction is stimulated by Co2+ and both reactions need Mg2+. The paralogues WbpW and PslB supply GDP-mannose for A-band LPS and the Psl exopolysaccharide.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000271 polysaccharide biosynthetic process | IEA GO_REF:0000002 | TOO GENERAL | Summary: InterPro2GO from the bifunctional PMI/GMP family signature. Reason: True but uninformative. Alginic acid biosynthetic process (GO:0042121), accepted here, is a descendant and entails it. Supporting Evidence: file:PSEAE/algA/algA-uniprot.txt Produces a precursor for alginate polymerization. |
| GO:0004475 mannose-1-phosphate guanylyltransferase (GTP) activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000283179 (PMI-GMP clade, 30 leaf IBAs in 26 taxa), seeded by AlgA itself (PseudoCAP IDA, PMID:1846611) and E. coli ManC (P24174, EcoCyc IDA). The self-seed is expected, not circular. Reason: The exact activity of the purified enzyme. The IBD stands on ManC alone and the node term suits the clade. Supporting Evidence: PMID:1846611 For the GMP forward reaction, apparent Km values of 20.5 microM and 29.5 microM for D-mannose 1-phosphate and GTP, respectively, were obtained from double reciprocal plots. |
| GO:0004475 mannose-1-phosphate guanylyltransferase (GTP) activity | IDA PMID:1846611 Purification and characterization of phosphomannose isomeras... | ACCEPT | Summary: PseudoCAP IDA; seed of the PTN000283179 GMP-activity IBD. Seed verdict SOUND. Reason: Purified PMI-GMP (N-terminally sequenced as AlgA in Swiss-Prot) catalyses the GMP reaction in both directions with measured kinetics. Supporting Evidence: PMID:1846611 For the GMP forward reaction, apparent Km values of 20.5 microM and 29.5 microM for D-mannose 1-phosphate and GTP, respectively, were obtained from double reciprocal plots. file:PSEAE/algA/algA-uniprot.txt KM=20.5 uM for D-mannose 1-phosphate |
| GO:0004475 mannose-1-phosphate guanylyltransferase (GTP) activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined IEA from the GMPP domain, Rhea and EC 2.7.7.13. Reason: Consistent with the purified-enzyme evidence. Supporting Evidence: PMID:1846611 For the GMP forward reaction, apparent Km values of 20.5 microM and 29.5 microM for D-mannose 1-phosphate and GTP, respectively, were obtained from double reciprocal plots. |
| GO:0004476 mannose-6-phosphate isomerase activity | IDA PMID:1846611 Purification and characterization of phosphomannose isomeras... | ACCEPT | Summary: PseudoCAP IDA for PMI activity of the purified enzyme. Reason: PMI activity co-purifies with GMP activity in a single monomeric protein, with measured Km and Vmax. Supporting Evidence: PMID:1846611 The apparent Km for D-mannose 6-phosphate was 3.03 mM, and the Vmax was 830 nmol/min/mg of enzyme. PMID:1846611 The PMI and GMP activities co-eluted in the same protein peak through successive fractionation on hydrophobic interaction, ion exchange, and gel filtration chromatography. |
| GO:0004476 mannose-6-phosphate isomerase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined IEA from Rhea and EC 5.3.1.8. Reason: Consistent with the purified-enzyme evidence. Supporting Evidence: PMID:1846611 The apparent Km for D-mannose 6-phosphate was 3.03 mM, and the Vmax was 830 nmol/min/mg of enzyme. |
| GO:0005976 polysaccharide metabolic process | IEA GO_REF:0000002 | TOO GENERAL | Summary: InterPro2GO from the type II PMI C-terminal domain. Reason: True but uninformative. Alginic acid biosynthetic process (GO:0042121), accepted here, is a descendant and entails it. Supporting Evidence: file:PSEAE/algA/algA-uniprot.txt Produces a precursor for alginate polymerization. |
| GO:0009298 GDP-mannose biosynthetic process | IBA GO_REF:0000033 | ACCEPT | Summary: IBA from PTN000283179, seeded by E. coli ManC (P24174, EcoCyc EXP) and the paralogue WbpW (Q9HTB7, PseudoCAP row from E. coli complementation; see the wbpW review for its evidence-code CHECK). Reason: AlgA catalyses two of the three steps from fructose 6-phosphate to GDP-mannose. The IBD is sound on ManC regardless of the WbpW evidence code. Supporting Evidence: PMID:1846611 We report here the purification and characterization of phosphomannose isomerase-guanosine 5'-diphospho-D-mannose pyrophosphorylase, a bifunctional enzyme (PMI-GMP) which catalyzes both the phosphomannose isomerase (PMI) and guanosine 5'-diphospho-D-mannose pyrophosphorylase (GMP) reactions of the Pseudomonas aeruginosa alginate biosynthetic pathway. |
| GO:0009298 GDP-mannose biosynthetic process | IEA GO_REF:0000041 | ACCEPT | Summary: UniPathway mapping (UPA00126, GDP-alpha-D-mannose biosynthesis). Reason: Consistent with the biochemistry; same action as for algC and wbpW. Supporting Evidence: PMID:1846611 We report here the purification and characterization of phosphomannose isomerase-guanosine 5'-diphospho-D-mannose pyrophosphorylase, a bifunctional enzyme (PMI-GMP) which catalyzes both the phosphomannose isomerase (PMI) and guanosine 5'-diphospho-D-mannose pyrophosphorylase (GMP) reactions of the Pseudomonas aeruginosa alginate biosynthetic pathway. |
| GO:0016779 nucleotidyltransferase activity | IEA GO_REF:0000002 | TOO GENERAL | Summary: InterPro2GO nucleotidyltransferase activity. Reason: True, but mannose-1-phosphate guanylyltransferase (GTP) activity (GO:0004475), accepted here, is a descendant and entails it. Supporting Evidence: PMID:1846611 For the GMP forward reaction, apparent Km values of 20.5 microM and 29.5 microM for D-mannose 1-phosphate and GTP, respectively, were obtained from double reciprocal plots. |
| GO:0042121 alginic acid biosynthetic process | IMP PMID:1846611 Purification and characterization of phosphomannose isomeras... | ACCEPT | Summary: PseudoCAP IMP from an algA point mutant with reduced PMI and GMP activity. Reason: AlgA makes the GDP-mannose that AlgD oxidises to the alginate donor GDP-mannuronic acid, so it performs early steps of alginate synthesis. The abstract reports the algA mutation that lowers both activities; the alginate phenotype is not stated in the abstract but is the basis of the original alg mutant, and the role is curated in Swiss-Prot from this paper. Same action as for algC. Supporting Evidence: PMID:1846611 DNA sequencing of a cloned mutant algA gene from P. aeruginosa revealed that a point mutation at nucleotide 961 greatly decreased the levels of both PMI and GMP in a crude extract. file:PSEAE/algA/algA-uniprot.txt Produces a precursor for alginate polymerization. |
Loading supporting content…
Download this section (compressed HTML)Q: How is GDP-mannose partitioned between the alginate (AlgA), A-band (WbpW) and Psl (PslB) pathways when the three paralogous PMI-GMPs are co-expressed?
Experiment: Measure intracellular GDP-mannose and alginate output in algA, wbpW and pslB single and combined deletions of a mucoid strain, to test whether the paralogues can substitute for AlgA in alginate synthesis.
Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)