Cloned (Comment) | Organism |
---|---|
DNA and amino acid sequence determination and analysis, recombinant expression of wild-type and truncation and point mutant enzymes in Escherichia coli strain BL21(DE3) using plasmid pSF49 | Streptococcus pyogenes phage H4489A |
Protein Variants | Comment | Organism |
---|---|---|
D170T | site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme | Streptococcus pyogenes phage H4489A |
additional information | construction of truncated mutants hylp255e1113 and hylp390e1113, HylP enzyme without internal Gly-X-Y motif | Streptococcus pyogenes phage H4489A |
Q295E | site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme | Streptococcus pyogenes phage H4489A |
W19F | site-directed mutagenesis, denaturation profile with guanidinium hydrochloride is similar to the wild-type enzyme | Streptococcus pyogenes phage H4489A |
Y182F | site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme | Streptococcus pyogenes phage H4489A |
Y298F | site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme | Streptococcus pyogenes phage H4489A |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
Ca2+ | interacts with the collagenous Gly-X-Y motif of the enzyme, activates up to 20 mM, but inhibits at higher concentrations, inactivation above 50 mM | Streptococcus pyogenes phage H4489A | |
L-ascorbate | noncompetitive inhibition, inhibition kinetics, overview. Residues involved in the binding of L-ascorbate are confined to HylP135-308 | Streptococcus pyogenes phage H4489A |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Streptococcus pyogenes phage H4489A | |
0.285 | - |
hyaluronic acid | pH 6.0, 25°C, recombinant wild-type enzyme | Streptococcus pyogenes phage H4489A | |
0.305 | - |
hyaluronic acid | pH 6.0, 25°C, recombinant mutant Y298F | Streptococcus pyogenes phage H4489A | |
0.373 | - |
hyaluronic acid | pH 6.0, 25°C, recombinant mutant Q295E | Streptococcus pyogenes phage H4489A |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Ca2+ | interacts with the collagenous Gly-X-Y motif of the enzyme, activates up to 20 mM, but inhibits at higher concentrations | Streptococcus pyogenes phage H4489A |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
40000 | - |
3 * 40000, SDS-PAGE | Streptococcus pyogenes phage H4489A |
120000 | - |
recombinant enzyme, glutaraldehyde cross-linking | Streptococcus pyogenes phage H4489A |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Streptococcus pyogenes phage H4489A | P15316 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant enzymes from Escherichia coli | Streptococcus pyogenes phage H4489A |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
hyaluronic acid | the enzyme is a hyaluronic acid specific hyaluronate lyase showing ability to degrade hyaluronate to unsaturated disaccharide units | Streptococcus pyogenes phage H4489A | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
More | extended trimeric conformation, homology modeling using the crystal structure of HylP1 from Streptococcus pyogenes SF370 (PDB ID-2C3F) as a template, domain structure, overview | Streptococcus pyogenes phage H4489A |
trimer | 3 * 40000, SDS-PAGE | Streptococcus pyogenes phage H4489A |
Synonyms | Comment | Organism |
---|---|---|
HylP | - |
Streptococcus pyogenes phage H4489A |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Streptococcus pyogenes phage H4489A |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
2.76 | - |
hyaluronic acid | pH 6.0, 25°C, recombinant mutant Q295E | Streptococcus pyogenes phage H4489A | |
4.29 | - |
hyaluronic acid | pH 6.0, 25°C, recombinant mutant Y298F | Streptococcus pyogenes phage H4489A | |
5.65 | - |
hyaluronic acid | pH 6.0, 25°C, recombinant wild-type enzyme | Streptococcus pyogenes phage H4489A |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
6 | - |
assay at | Streptococcus pyogenes phage H4489A |
IC50 Value | IC50 Value Maximum | Comment | Organism | Inhibitor | Structure |
---|---|---|---|---|---|
0.7 | - |
pH 6.0, 25°C, recombinant wild-type enzyme | Streptococcus pyogenes phage H4489A | L-ascorbate |
General Information | Comment | Organism |
---|---|---|
malfunction | the enzyme activity is considerably reduced with mutation in the second pocket consisting of Glu295 and Tyr298 | Streptococcus pyogenes phage H4489A |
additional information | The primary catalytic residues of the enzyme seem to be in the first pocket consisting of Asp170 and Tyr182. Catalytic residues span between the regions containing 135-308 amino acids where both the catalytic pocket has a prominent positively charged residue, structure overview. The collagenous Gly-X-Y motif of the enzyme influences stability and interact with calcium ions suggesting its role in the enzyme regulation | Streptococcus pyogenes phage H4489A |
physiological function | hyaluronate lyase is proposed to be one of the key bacteriophage-encoded virulence factors. The Gly-X-Y motif of HylP has a regulatory role for enzyme function | Streptococcus pyogenes phage H4489A |