Alpha-crystallin promotes assembly of a trimeric form of Mycobacterium tuberculosis Hsp16.3 in a cell free system.
Hsp16.3, a small heat shock protein from Mycobacterium tuberculosis proposed to form specific trimer-of-trimers structures, acts as a molecular chaperone in vitro. The assembly mechanisms of this oligomeric protein were studied using in vitro transcription/translation systems. Analysis using a combination of non-denaturing pore gradient polyacrylamide gel electrophoresis and size-exclusion chromatography demonstrates that the predominant form of Hsp16.3 produced in the in vitro transcription/translation system is the trimer. Our result indicated that alpha-crystallin (molecular chaperone) remarkably promotes the trimer assembly of Hsp16.3, but does not convert the trimeric form to nonameric form. An "inert" Hsp16.3 dimer, which does not seem to participate in trimer assembly but may be involved in forming other forms of Hsp16.3, was also detected in the in vitro expression system. A latent phase of ~10 min in the appearance of the first detectable species indicated that Hsp16.3 assembly did not occur co-translationally.
Abulimiti
A
A
Department of Biological Science and Biotechnology, School of Life Science, Tsinghua University, The Protein Science Laboratory of the Education Ministry, Beijing, 100084 P. R. China.
Chang
Z
Z
D001426:Bacterial Proteins; D001681:Biological Assay; D002474:Cell-Free System; D018833:Chaperonins / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D002850:Chromatography, Gel; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004926:Escherichia coli / Q000378:metabolism; D007700:Kinetics; D009169:Mycobacterium tuberculosis / Q000737:chemistry* / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D014176:Protein Biosynthesis; D011487:Protein Conformation; D014357:Trypsin / Q000378:metabolism; D038201:alpha-Crystallins / Q000737:chemistry* / Q000494:pharmacology
D001426:Bacterial Proteins; C431133:Hsp16.3 protein, Mycobacterium tuberculosis; D038201:alpha-Crystallins; D014357:Trypsin; D018833:Chaperonins