Engineering enzymes for stability

Current Opinion in Structural Biology

Volumn 6, Issue 4, 1996, Pages 546-550

  Shaw, Andrew ^a   Bott, Richard ^a  

^a GENENCOR INTERNATIONAL INC   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

ENZYME DENATURATION; ENZYME STABILITY; GENETIC ENGINEERING; KINETICS; PRIORITY JOURNAL; PROTEIN FOLDING; REVIEW; THERMODYNAMICS;

EID: 0030220763     PISSN: 0959440X     EISSN: None     Source Type: Journal    
DOI: 10.1016/S0959-440X(96)80122-9     Document Type: Article

References (45)

1. of special interest Zhu H, Dupureur CM, Zhang X, Tsai M-D. Phospholipase A2 engineering. the roles of disulfide bonds in structure, conformational stability and catalytic function. Biochemistry. 34:1995;15307-15314 Each of seven disulfide bridges in the 123 amino acid PLA2 have been replaced pairwise by the mutation of Cys→Ala cysala. NMR structural analysis of the six folded variants found that although the variants retained enzymatic activity, many were in less-ordered states than the native enzyme.
2. of special interest Itzhaki LS, Otzen DE, Fersht AR. The structure of the transition state for folding of chymotrypsin inhibitor 2 analysed by protein engineering methods: evidence for a nucleation - condensation mechanism for protein folding. J Mol Biol. 245:1995;260-288 A very concise analysis of numerous variants of CI2 that have been analyzed for their relative conformational stability and alterations in their folding and unfolding kinetics. The focus of the paper is to identify residues that are already folded in the transition state and thus can be used to define the structure of the transition state. The finding is that residues in the N-terminal helix may already fold in the transition state. For the purposes of this review, the finding that most are not folded in the transition state is more relavant.
3. of special interest Sosnick TS, Jackson S, Wilk RR, Englander SW, DeGrado WF. The role of helix formation in the folding of a fully α-helical coiled coil. Proteins. 24:1996;427-432 An analysis of the folding and dimerization of GCN4. The very concise discussion of the relevance of folding kinetics to the analysis of transition states is well worth reading.
4. of special interest Milla ME, Brown BM, Waldburger CD, Sauer RT. P22 arc repressor: transition state properties inferred from mutational effects on the rates of protein unfolding and refolding. Biochemistry. 34:1995;13914-13919 Another analysis of folding kinetics and its relevance to stability. This study, taken in conjunction with [2] and [3] makes a compelling statement of the limited amount of ordered structure in the folding transition states of these simple folding systems.
5. Matthews BW. Structural and genetic analysis of protein stability. Annu Rev Biochem. 62:1993;139-160.
6. of outstanding interest Shortle D. The denatured state (the other half of the folding equation) and its role in protein stability. FASEB J. 10:1996;27-34 This is an excellent review, examining stability from the point of view of the relative stability of the unfolded state. Taken in conjunction with the references cited within, this review constitutes a must read for all who want to stabilize folded proteins.
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