Role of Active Site Rigidity in Activity: MD Simulation and Fluorescence Study on a Lipase Mutant

Relationship between stability and activity of enzymes is maintained by underlying conformational flexibility. In thermophilic enzymes, a decrease in flexibility causes low enzyme activity while in less stable proteins such as mesophiles and psychrophiles, an increase in flexibility is associated with enhanced enzyme activity. Recently, we identified a mutant of a lipase whose stability and activity were enhanced simultaneously. In this work, we probed the conformational dynamics of the mutant and the wild type lipase, particularly flexibility of their active site using molecular dynamic simulations and time-resolved fluorescence techniques. In contrast to the earlier observations, our data show that active site of the mutant is more rigid than wild type enzyme. Further investigation suggests that this lipase needs minimal reorganization/flexibility of active site residues during its catalytic cycle. Molecular dynamic simulations suggest that catalytically competent active site geometry of the mutant is relatively more preserved than wild type lipase, which might have led to its higher enzyme activity. Our study implies that widely accepted positive correlation between conformation flexibility and enzyme activity need not be stringent and draws attention to the possibility that high enzyme activity can still be accomplished in a rigid active site and stable protein structures. This finding has a significant implication towards better understanding of involvement of dynamic motions in enzyme catalysis and enzyme engineering through mutations in active site

Sepsis: in search of cure

Introduction Sepsis is a complex inflammatory disorder believed to originate from an infection by any types of microbes and/or their products. It is the leading cause of death in intensive care units (ICUs) throughout the globe.The mortality rates depend both on the severity of infection and the host’s response to infection.Methods Literature survey on pathobiology of sepsis in general and failure of more than hundred clinical trialsconducted so far in search of a possible cure for sepsisresulted in the preparation of this manuscript.FindingsSepsis lacks a suitable animal model thatmimics human sepsis. However, based on the resultsobtained in animal models of sepsis, clinical trials con-ducted so far have been disappointing. Althoughinvolvement of multiple mediators and pathways in sepsishas been recognized, only few components are being tar-geted and this could be the major reason behind the failureof clinical trials.ConclusionInability to recognize a single critical medi-ator of sepsis may be the underlying cause for the poortherapeutic intervention of sepsis. Therefore, sepsis is stillconsidered as a disease—in search of cu