Expression and purification of a protein kinase, PknL of Mycobacterium bovis BCG pasteur 1173P2

Lin, Li Shin (2005) Expression and purification of a protein kinase, PknL of Mycobacterium bovis BCG pasteur 1173P2. Universiti Malaysia Sabah. (Unpublished)


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Signal transduction pathways in both eukaryotes and prokaryotes involve specific protein kinases coupled to protein phosphatase. Genome sequence of Mycobacterium tuberculosis H37Rv revealed the existence of eleven eukaryotic Serrrbr protein kinases (STPK). In this project, pknL, which is one of STPK of Mycobacterium bovis BeG Pasteur 1173P2 was inserted into expression vector, pET-16b which was then transformed into expression host, E.coli BL21 (DE3) pLysS. IPTG was used to induce pknL gene expression followed by recombinant protein purification with Ni-NT A Spin Kit. The molecular weight of purified recombinant PknL was determined using SDSPAGE. Recombinant PknL has molecular weight of 45 kDa. Most of recombinant protein was purified in first elution while remaining proteins purified in second elution. Full amino acids sequence of pknL obtained from database was subjected to bioinfonnatics analysis. Analysis of the amino acid sequence of PknL with Dense Alignment Surface (DAS) and PHD predictions indicated the presence of transmembrane domain at amino acid 369 - 389 within the helical C-terminal ofPknL. Protein kinase domain was found at amino acid 27 - 286 at the N-terminal and conserved kinase domain was showed to be 99% similar to corresponding kinases in 'Mycobacterium tuberculosis H37Rv and Mycobacterium bovis AF2122/97. Further analysis revealed that the secondary structure of PknL comprised mainly of helices and loops with a helices transmembrane structure and loopy structure at Serrrbr protein kinase active site. These finding showed that PknL is a membrane spanning protein in Mycobacterium bovis BCG Pasteur 1173P2.

Item Type: Academic Exercise
Uncontrolled Keywords: Genome, Mycobacterium tuberculosis, Bioinformatics analysis, Protein kinase, Pasteur
Subjects: Q Science > QP Physiology > QP1-(981) Physiology > QP501-801 Animal biochemistry
Divisions: SCHOOL > School of Science and Technology
Depositing User: ADMIN ADMIN
Date Deposited: 13 Jul 2012 10:33
Last Modified: 23 Oct 2017 12:41

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