Rescue of deleterious mutations by the compensatory Y30F mutation in ketosteroid isomerase
SCIE
SCOPUS
KCI
- Title
- Rescue of deleterious mutations by the compensatory Y30F mutation in ketosteroid isomerase
- Authors
- Hyung Jin Cha; Doo Soo Jang; Yeon -gil Kim; Bee- Hak Hong; Jae-sung Woo; Kim, KT; Choi, KY
- Date Issued
- 2013-07
- Publisher
- KSBMB
- Abstract
- Proteins have evolved to compensate for detrimental mutations. However, compensatory mechanisms for protein defects are not well understood. Using ketosteroid isomerase (KSI), we investigated how second-site mutations could recover defective mutant function and stability. Previous results revealed that the Y30F mutation rescued the Y14F, Y55F and Y14F/Y55F mutants by increasing the catalytic activity by 23-, 3- and 1.3-fold, respectively, and the Y55F mutant by increasing the stability by 3.3 kcal/mol. To better understand these observations, we systematically investigated detailed structural and thermodynamic effects of the Y30F mutation on these mutants. Crystal structures of the Y14F/Y30F and Y14F/Y55F mutants were solved at 2.0 and 1.8 previoulsy solved structures of wild-type and other mutant KSIs. Structural analyses revealed that the Y30F mutation partially restored the active-site cleft of these mutant KSIs. The Y30F mutation also increased Y14F and Y14F/Y55F mutant stability by 3.2 and 4.3 kcal/mol, respectively, and the melting temperatures of the Y14F, Y55F and Y14F/Y55F mutants by 6.4A degrees C, 5.1A degrees C and 10.0A degrees C, respectively. Compensatory effects of the Y30F mutation on stability might be due to improved hydrophobic interactions because removal of a hydroxyl group from Tyr30 induced local compaction by neighboring residue movement and enhanced interactions with surrounding hydrophobic residues in the active site. Taken together, our results suggest that perturbed active-site geometry recovery and favorable hydrophobic interactions mediate the role of Y30F as a secondsite suppressor.
- Keywords
- active-site recovery; ketosteroid isomerase; more hydrophobic interactions; rescue mechanism; second-site suppressor; HYDROGEN-BOND NETWORK; PUTIDA BIOTYPE-B; ONCOGENIC P53 MUTATIONS; DELTA(5)-3-KETOSTEROID ISOMERASE; ACTIVE-SITE; SUPPRESSOR MUTATIONS; T4 LYSOZYME; CRYSTAL-STRUCTURE; STRUCTURAL BASIS; DOUBLE-MUTANT
- URI
- https://oasis.postech.ac.kr/handle/2014.oak/15014
- DOI
- 10.1007/S10059-013-0013-1
- ISSN
- 1016-8478
- Article Type
- Article
- Citation
- Molecules and cells, vol. 36, no. 1, page. 39 - 46, 2013-07
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