An integrated platform for automatic design and screening of virtual mutants based on 3D-QSAR analysis

• The work represents the first example of 3D-QSAR strategies applied to enzyme rational engineering.
• The computational methodology allows for rational protein engineering using a very moderate computational power.
• 3D-QSAR paves the way for a rational exploitation of large database of enzyme structures thus making possible the transformation of data into usable knowledge.
• The application of the automatic work-flow for software integration is a novel tool for highthroughput screening of virtual mutants.

An innovative application of 3D-QSAR methodology to the rational design of enzymes is here reported. The introduction of amidase activity inside the scaffold of lipase B from Candida antarctica (CaLB) was studied and 3D-QSAR models were constructed to correlate the structures of a set of CaLB mutants with their experimentally measured activities. Properties, like hydrophilicity, hydrophobicity and hydrogen bonding capability of the enzyme active site were computed by means of the GRID method and the output was used as molecular descriptors. Correlations with experimental behavior of the catalysts were calculated by means of partial least square regression (PLS). The analysis of the QSAR model fully exploits fundamental knowledge while avoiding conceptual biases. Rationales for driving enzyme engineering are disclosed and a priori evaluation of new virtual candidate mutants becomes feasible. On that respect, the whole procedure for production of virtual mutants and scoring of their activity was automated within a workflow constructed by means of the modeFRONTIER package. The method allows for the automated construction and scoring of each mutant in 2 h on a normal workstation.

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