Detail of GM0013


Summary

The genetically modified enzybiotic, named Ply187AN-KSH3b , constructed by Domains Assembly strategy, improve the lytic activity.
With  Ply187AN-KSH3b
  Sequence Length:  225 AA.
  Mass:  25079.1 Da.
  Isoelectric Point:  9.33
  Function:  The fusion’s MIC is five- fold lower than that of LysK’s (and twofold lower when compared on a molar basis)

Construction

GM0013, constructed by Domains Assembly strategy.
O56785: Cell wall hydrolase Ply187
I6X5A8: LysK
UniProt

IPR007921:11-145
IPR007921:14-145
IPR013338:476-622
IPR013338:476-622
IPR007921:29-160
IPR002502:196-369
IPR003646:409-481
InterPro

GM0013:1-157
GM0013:158-293
GMEnzy

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Details:
GM0013[1-157]: derive from protein O56785 sequence 1-157

O56785, 628 AA., the Cell wall hydrolase Ply187 from Staphylococcus phage 187

Source: Staphylococcus phage 187

Domains and repeats

11-145:  IPR007921, the CHAP domain
476-622:  IPR013338, the Lysozyme domain, subfamily 2
14-145:  IPR007921, the CHAP domain
476-622:  IPR013338, the Lysozyme domain, subfamily 2

GO term prediction

Biological Process:  0009253, the peptidoglycan catabolic process
Biological Process:  0044036, the cell wall macromolecule metabolic process
Molecular Function:  0004040, the amidase activity?
Molecular Function:  0016787, the hydrolase activity?
Biological Process:  0009253, the peptidoglycan catabolic process
Biological Process:  0044036, the cell wall macromolecule metabolic process?
Molecular Function:  0004040, the amidase activity?
Molecular Function:  0016787, the hydrolase activity?


Linking to UniprotKB
Linging to InterPro
GM0013[158-293]: derive from protein I6X5A8 sequence 409-481

I6X5A8, 495 AA., the LysK from Staphylococcus phage Fi200W

Source: Staphylococcus phage Fi200W

Domains and repeats

29-160:  IPR007921, the CHAP domain
196-369:  IPR002502, the N-acetylmuramoyl-L-alanine amidase domain
409-481:  IPR003646, the SH3-like domain, bacterial-type?

GO term prediction

Molecular Function:  0008745, the N-acetylmuramoyl-L-alanine amidase activity


Linking to UniprotKB
Linging to InterPro

Annotation

1-157
158-293
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1123

1-157
: EAD, CHAP domain, derived from S. aureus bacteriophage 187 endolysin;
158-293
: CBD, SH3b domain, derived from anti-staphylococcal bacteriophage endolysin, LysK;
Domain and repeats:
11-145
: IPR007921,CHAP domain;
14-145
: IPR007921,CHAP domain;
158-293
: IPR003646,SH3-like domain, bacterial-type?;
GO term prediction:
MALPKTGKPTAKQVVDWAINLIGSGVDVDGYYGRQCWDLPNYIFNRYWNFKTPGNARDMAWYRYPEGFKVFRNTSDFVPKPGDIAVWTGGNYNWNTWGHTGIVVGPSTKSYFYSVDQNWNNSNSYVGSPAAKIKHSYFGVTHFVRPAYKAEPKPTP-ENATFVNGNQPIVTRIGSPFLNAPVGGNLPAGATIVYDEVCIQAGHIWIGYNAYNGNRVYCPVRTCQGV
  BlastP to GMEnzy

Production

  1.  Expressed by pET21a in E. coli BL21(DE3),  Purified by nickel chromatography Ni-NTA (Qiagen, Valencia, CA)

Activity

  1.  MIC test on Staphylococcus aureus (S. aureus Newman) by microdilution broth method
  showed 7.6±2.9μg/mL(0.24±0.09 nmol/mL)

Reference

  1. Mao, J.Schmelcher, M.Harty, W. J.Foster-Frey, J.Donovan, D. M.. (2013) Chimeric Ply187 endolysin kills Staphylococcus aureus more effectively than the parental enzyme. FEMS Microbiol Lett. 01:30-36. [doi:10.1111/1574-6968.12104] [PMID:23413880] [FULL TEXT]

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