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Title 

Contribution of individual side-chains to the stability of BPTI examined by alanine-scanning mutagenesis

Authors 

Myeong Hee YuJ WeissmanPS Kim

Publisher 

Elsevier

Issue Date 

1995

Citation 

Journal of Molecular Biology, vol. 249, no. 0, pp. 388-397

Keywords 

BPTIprotein foldingprotein stabilityalaninecalorimetry, differential scanningmutagenesis, site-directed

Abstract 

Bovine pancreatic trypsin inhibitor (BPTI) serves as an important model system for the examination of almost all aspects of protein structure. Systematic studies of the effects of mutation on the thermodynamic stability of BPTI, however, have been limited by the extreme stability of the protein. A derivative of BPTI containing only the 5-55 disulfide bend, termed [5-55](Ala), has been shown previously to fold into a structure wry similar to that of native BPTI and to be a functional trypsin inhibitor. [5-55](Ala) undergoes a reversible thermal unfolding transition with a melting temperature of 39°C, and is therefore well suited for stability studies. Using an alanine-scanning mutagenesis approach, we have examined the contribution to stability of each side-chain in the [5-55](Ala) derivative of BPTI. These studies demonstrate the importance of the two hydrophobic cores composed largely of clusters of aromatic residues, as well as the internal hydrogen-bonding network, in stabilizing BPTI. Overall, there is a strong relationship between change in buried surface area and stability for both polar and hydrophobic residues, with proportionality constants of 50 and 20 cal/?2, respectively None of the alanine substitutions substantially stabilized [5-55](A?). Nonetheless, approximately 60% (28/46) of the alanine mutants were destabilized by less than 10°C, suggesting that a form of BPTI with up to half of its residues being alanine could fold into a stable structure resembling the native one.

ISSN 

0022-2836

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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