In Vitro Holdase Activity of E. coli Small Heat-Shock Proteins IbpA, IbpB and IbpAB: A Biophysical Study with Some Unconventional Techniques
E. coli small heat shock proteins IbpA and IbpB (inclusion body binding proteins A and B) are known to act as holding chaperones on denaturing, aggregate-prone proteins. But, there is no clear understanding about which of the IbpA and IbpB has more holdase activity and how the holdase
activity of one was influenced by the presence of the other. This study was conducted to resolve the questions, using some uncommon physical techniques like dynamic light scattering, micro-viscometry and atomic force microscopy in addition to the common techniques of spectrophotometry and
spectrofluorimetry. The holdase activity was investigated on the heat-denatured L-lactate dehydrogenase (LDH) of rabbit muscle. LDH was found to be deactivated completely without any aggregation at 52°C and with transient aggregation at 60°C; molecular dynamics simulation also revealed
that at 52°C, denaturation occurred only at the active site of LDH. When LDH was allowed to be deactivated in the presence of IbpA, IbpB or (IbpA + IbpB), partial inhibition of i) denaturation at 52°C and ii) aggregation at 60°C were observed. The results further demonstrated that
the holdase activity of IbpB was higher than that of IbpA and their combined effect was higher than their individual one.
Keywords: Atomic force microscopy; E. coli Small heat shock proteins IbpA & IbpB; dynamic light scattering; holdase activity; lactate dehydrogenase; micro-viscometry
Document Type: Research Article
Publication date: 01 June 2014
- Protein & Peptide Letters publishes short papers in all important aspects of protein and peptide research, including structural studies, recombinant expression, function, synthesis, enzymology, immunology, molecular modeling, drug design etc. Manuscripts must have a significant element of novelty, timeliness and urgency that merit rapid publication. Reports of crystallisation, and preliminary structure determinations of biologically important proteins are acceptable. Purely theoretical papers are also acceptable provided they provide new insight into the principles of protein/peptide structure and function.
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