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Literature summary for 2.4.1.212 extracted from

  • Kooy, F.; Beeftink, H.; Eppink, M.; Tramper, J.; Eggink, G.; Boeriu, C.
    Kinetic and structural analysis of two transferase domains in Pasteurella multocida hyaluronan synthase (2014), J. Mol. Catal. B, 102, 138-145.
No PubMed abstract available
Search for more literature for 2.4.1.212

Cloned(Commentary)

Cloned (Comment) Organism
gene hyaD, recombinant expression of the soluble C-terminally His6-tagged PmHAS1-703 truncated enzyme from pET101/D-TOPO expression vector with an additional V5 epitope Pasteurella multocida

Inhibitors

Inhibitors Comment Organism Structure
UMP dead-end inhibitor Pasteurella multocida

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information structural and kinetic analysis and modeling, both NAc- and UA-transferase domains follow a sequential kinetic mechanism, most likely an ordered one in which the UDP-sugar donor binds first, followed by the HA oligosaccharide. After transfer of the sugar moiety, both products are released, first the elongated HA oligosaccharide and then the UDP sugar. A mechanistic shift from a steady-state ordered bi-bi to rapid equilibrium ordered bi-bi mechanism is observed at the NAc-site between the HA6 and HA8 elongation, detailed overview Pasteurella multocida
0.6
-
 hyaluronan oligomer HA5 with UDP-alpha-D-glucuronate, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida
0.7
-
 hyaluronan oligomer HA4 with UDP-alpha-N-acetyl-D-glucosamine, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida
0.7
-
 hyaluronan oligomer HA8 with UDP-alpha-N-acetyl-D-glucosamine, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida
1
-
 hyaluronan oligomer HA6 with UDP-alpha-N-acetyl-D-glucosamine, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Pasteurella multocida

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
additional information Pasteurella multocida Pasteurella multocida hyaluronan synthase encompasses two transferase domains that elongate a growing hyaluronan oligosaccharide chain by addition of either GlcNAc or GlcUA residues from a corresponding UDP-sugar ?
-
 ?
UDP-alpha-D-glucuronate + N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-[nascent hyaluronan] Pasteurella multocida
-
 UDP + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-[nascent hyaluronan]
-
 ?
UDP-alpha-N-acetyl-D-glucosamine + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-[nascent hyaluronan] Pasteurella multocida
-
 UDP + N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-[nascent hyaluronan]
-
 ?

Organism

Organism UniProt Comment Textmining
Pasteurella multocida Q7BLV3 gene hyaD
-

Purification (Commentary)

Purification (Comment) Organism
recombinant soluble C-terminally His6-tagged PmHAS1-703 by nickel affinity chromatography Pasteurella multocida

Reaction

Reaction Comment Organism Reaction ID
UDP-alpha-D-glucuronate + N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-[nascent hyaluronan] = UDP + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-[nascent hyaluronan] a mechanistic shift from a steady-state ordered bi-bi to rapid equilibrium ordered bi-bi mechanism is observed at the NAc-site between the HA6 and HA8 elongation Pasteurella multocida
a
UDP-N-acetyl-alpha-D-glucosamine + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-[nascent hyaluronan] = UDP + N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-[nascent hyaluronan] a mechanistic shift from a steady-state ordered bi-bi to rapid equilibrium ordered bi-bi mechanism is observed at the NAc-site between the HA6 and HA8 elongation Pasteurella multocida
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
additional information Pasteurella multocida hyaluronan synthase encompasses two transferase domains that elongate a growing hyaluronan oligosaccharide chain by addition of either GlcNAc or GlcUA residues from a corresponding UDP-sugar Pasteurella multocida ?
-
 ?
additional information initial velocity studies of single-step elongations are conducted for both domains by independently varying the concentrations of the hyaluronan oligosaccharide and the UDP-sugar. Two-substrate models are discriminated by their goodness-of-fit parameters and by dead-end inhibition studies. Coupled-enzyme assay using LDH, PK; NADH and phosphoenolpyruvate, as well as hyaluronan oligosaccharides, UDP-GlcNAc and UDP-GlcUA, overview Pasteurella multocida ?
-
 ?
UDP-alpha-D-glucuronate + hyaluronan oligomer HA5
-
 Pasteurella multocida UDP + beta-D-glucuronosyl-(1->3)-hyaluronan oligomer HA5
-
 ?
UDP-alpha-D-glucuronate + N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-[nascent hyaluronan]
-
 Pasteurella multocida UDP + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-[nascent hyaluronan]
-
 ?
UDP-alpha-N-acetyl-D-glucosamine + beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-[nascent hyaluronan]
-
 Pasteurella multocida UDP + N-acetyl-beta-D-glucosaminyl-(1->4)-beta-D-glucuronosyl-(1->3)-N-acetyl-beta-D-glucosaminyl-(1->4)-[nascent hyaluronan]
-
 ?
UDP-alpha-N-acetyl-D-glucosamine + hyaluronan oligomer HA4
-
 Pasteurella multocida UDP + N-acetyl-beta-D-glucosaminyl-(1->4)-hyaluronan oligomer HA4
-
 ?
UDP-alpha-N-acetyl-D-glucosamine + hyaluronan oligomer HA6
-
 Pasteurella multocida UDP + N-acetyl-beta-D-glucosaminyl-(1->4)-hyaluronan oligomer HA6
-
 ?
UDP-alpha-N-acetyl-D-glucosamine + hyaluronan oligomer HA8
-
 Pasteurella multocida UDP + N-acetyl-beta-D-glucosaminyl-(1->4)-hyaluronan oligomer HA8
-
 ?

Synonyms

Synonyms Comment Organism
PmHAS
-
 Pasteurella multocida

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
35
-
 assay at Pasteurella multocida

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
7.9
-
 hyaluronan oligomer HA5 with UDP-alpha-D-glucuronate, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida
7.9
-
 hyaluronan oligomer HA4 with UDP-alpha-N-acetyl-D-glucosamine, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida
8.8
-
 hyaluronan oligomer HA8 with UDP-alpha-N-acetyl-D-glucosamine, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida
13.5
-
 hyaluronan oligomer HA6 with UDP-alpha-N-acetyl-D-glucosamine, pH 8.0, 35°C, recombinant enzyme Pasteurella multocida

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
 assay at Pasteurella multocida