1T2N

Structure of a thermostable triple mutant of Bacillus subtilis lipase obtained through directed evolution

Summary for 1T2N

• Entry DOI: 10.2210/pdb1t2n/pdb
• Related: 1T4M
• Descriptor: Lipase, POTASSIUM ION (3 entities in total)
• Functional Keywords: alpha/beta hydrolase, hydrolase
• Biological source: Bacillus subtilis
• Cellular location: Secreted: P37957
• Total number of polymer chains: 1
• Total formula weight: 19498.97

Authors

Rajakumara, E.,Sankaranarayanan, R. (deposition date: 2004-04-22, release date: 2004-11-23, Last modification date: 2023-08-23)

Primary citation

Acharya, P.,Rajakumara, E.,Sankaranarayanan, R.,Rao, N.M.
Structural basis of selection and thermostability of laboratory evolved Bacillus subtilis lipase.
J.Mol.Biol., 341:1271-1281, 2004

PubMed Abstract: Variation in gene sequences generated by directed evolution approaches often does not assure a minimalist design for obtaining a desired property in proteins. While screening for enhanced thermostability, structural information was utilized in selecting mutations that are generated by error-prone PCR. By this approach we have increased the half-life of denaturation by 300-fold compared to the wild-type Bacillus subtilis lipase through three point mutations generated by only two cycles of error-prone PCR. At lower temperatures the activity parameters of the thermostable mutants are unaltered. High-resolution crystal structures of the mutants show subtle changes, which include stacking of tyrosine residues, peptide plane flipping and a better anchoring of the terminus, that challenge rational design and explain the structural basis for enhanced thermostability. The approach may offer an efficient and minimalist solution for the enhancement of a desired property of a protein.

PubMed: 15321721
DOI: 10.1016/j.jmb.2004.06.059

Experimental method

X-RAY DIFFRACTION (1.8 Å)