a 2011

Cloning, expression and structure determination of SH3b cell wall binding domain of bacteriophage 812 endolysin

BENEŠÍK, Martin, Jiří NOVÁČEK, Lubomír JANDA, Radka DOPITOVÁ, Lukáš ŽÍDEK et. al.

Basic information

Original name

Cloning, expression and structure determination of SH3b cell wall binding domain of bacteriophage 812 endolysin

Authors

BENEŠÍK, Martin (203 Czech Republic, belonging to the institution), Jiří NOVÁČEK (203 Czech Republic, belonging to the institution), Lubomír JANDA (203 Czech Republic, guarantor, belonging to the institution), Radka DOPITOVÁ (203 Czech Republic, belonging to the institution), Lukáš ŽÍDEK (203 Czech Republic, belonging to the institution), Jiří DOŠKAŘ (203 Czech Republic, belonging to the institution), Vladislava RŮŽIČKOVÁ (203 Czech Republic, belonging to the institution), Marek MOŠA (203 Czech Republic) and Roman PANTŮČEK (203 Czech Republic, belonging to the institution)

Edition

Phages 2011 - Bacteriophage Applications, 19-21 September 2011, St Hilda's College, Oxford, UK, 2011

Other information

Language

English

Type of outcome

Konferenční abstrakt

Field of Study

Genetics and molecular biology

Country of publisher

United Kingdom of Great Britain and Northern Ireland

Confidentiality degree

není předmětem státního či obchodního tajemství

References:

RIV identification code

RIV/00216224:14310/11:00050057

Organization unit

Faculty of Science

Keywords in English

bacteriophage therapy; Staphylococcus aureus; bacteriophage endolysin; SH3b domain

Tags

International impact, Reviewed
Změněno: 14/10/2011 12:57, prof. RNDr. Roman Pantůček, Ph.D.

Abstract

V originále

In this study we focused on endolysin of staphylococcal bacteriophage 812, a member of unclassified SPO1-like viruses from family Myoviridae. The endolysin of phage 812 includes three domains: two catalytic domains and the SH3b-domain that is probably a cell wall targeting domain, which could be responsible for binding of endolysin to peptidoglycan. The gene sequence for the C-terminal SH3b domain was cloned in pET28 vector and expressed as a soluble protein in E. coli BL21 (DE3) to determine the 3D structure of the protein. A native variant of the SH3b protein was purified without the his-tag. Subsequently, three variants of this protein have been prepared: (i) non-labeled, (ii) single-labeled (15N) and (iii) double-labeled (13C, 15N) domain. The protein was purified to homogeneity using ammonium sulphate precipitation, anion exchange chromatography, cation exchange chromatography, and gel filtration. Functionality of the cell wall binding of the purified protein has been verified by a co-sedimentation test in using S. aureus peptidoglycan.

Links

GAP206/11/0758, research and development project
Name: Vývoj metodologie s vysokým rozlišením pro NMR studie neuspořádaných proteinů s vysoce degenerovanými rezonančními frekvencemi (Acronym: HIGHRES)
Investor: Czech Science Foundation
GA310/09/0459, research and development project
Name: Úloha bakteriofágů v horizontálním přenosu genů virulence a rezistence u Staphylococcus aureus
Investor: Czech Science Foundation, Role of bacteriophages in horizontal transfer of virulence and drug resistance genes in Staphylococcus aureus
GP204/02/D099, research and development project
Name: Knihovna ribotypizačních profilů typových kultur bakterií
Investor: Czech Science Foundation, Database of the ribotype profiles of bacterial type strains
MSM0021622415, plan (intention)
Name: Molekulární podstata buněčných a tkáňových regulací
Investor: Ministry of Education, Youth and Sports of the CR, Molecular basis of cell and tissue regulations
TA01010405, research and development project
Name: Výzkum stafylokokových bakteriofágových mutant s širokým spektrem hostitelů (Acronym: TAČR/IMUNA-1)
Investor: Technology Agency of the Czech Republic