Effects of Distal Mutations on the Structure, Dynamics and Catalysis of Human Monoacylglycerol Lipase

Abstract

An understanding of how conformational dynamics modulates function and catalysis of human monoacylglycerol lipase (hMGL), an important pharmaceutical target, can facilitate the development of novel ligands with potential therapeutic value. Here, we report the discovery and characterization of an allosteric, regulatory hMGL site comprised of residues Trp-289 and Leu-232 that reside over 18 Å away from the catalytic triad. These residues were identified as critical mediators of long-range communication and as important contributors to the integrity of the hMGL structure. Nonconservative replacements of Trp-289 or Leu-232 triggered concerted motions of structurally distinct regions with a significant conformational shift toward inactive states and dramatic loss in catalytic efficiency of the enzyme. Using a multimethod approach, we show that the dynamically relevant Trp-289 and Leu-232 residues serve as communication hubs within an allosteric protein network that controls signal propagation to the active site, and thus, regulates active-inactive interconversion of hMGL. Our findings provide new insights into the mechanism of allosteric regulation of lipase activity, in general, and may provide alternative drug design possibilities.

Towards functional de novo designed proteins

“…That said, a design rule to emerge from this study is that certain charged side chains near the termini of helices stabilise the constructs, which is in agreement with conclusions drawn from a previous study combining bioinformatics and rational design of single a helices [85]. [86]. Therefore, design strategies that incorporate, or at least consider, the functional aspect at an early stage could ultimately lead to more successful outcomes.…”

A Thermolabile Phospholipase B from Talaromyces marneffei GD-0079: Biochemical Characterization and Structure Dynamics Study

“…The raw data at a given wavelength, λ (nm) were converted into concentration-independent parameter [θ] λ (deg cm is the mean residue weight of the protein, l is the path length of the cell in centimeters, and c is the protein concentration in mg ml −1 ). Baseline was corrected and smoothed (within permissible limits) by using the inbuilt CDNN software (Applied Photo physics, Surrey, UK) of the unit [16][17][18]. .…”

“…Circular Dichroism were performed for structural conformation and functional responses of a protein. Tyukhtenko et al, 2018 showed that CD spectra is indicative of the secondary structure prediction and no conformational change or folding may occurs which might be possible due to nonconservative substitutions at certain at certain length of spectra. Anwer et al, 2016 reported that secondary structural changes of a protein can be assessed from CD spectra and the protein follow two state folding mechanism.…”

“…Anwer et al, 2016 reported that secondary structural changes of a protein can be assessed from CD spectra and the protein follow two state folding mechanism. A lack of fixed tertiary interactions could results in a conformational shift towards ensemble of fluctuating structures [16][17][18]. The CD was performed in the far-UV region (190-260 nm) to predict the secondary structure and stability of TmPLB1.…”

Conformational gating, dynamics and allostery in human monoacylglycerol lipase

“…The molecular details of allosteric communications between the active site and the distal regions are still poorly understood. We have shown previously that the native state of hMGL is a conformational ensemble and that introducing an allosteric perturbation by single mutation of a critical distal residue can significantly affect the networks of allosteric interactions, resulting in conformational rearrangements in the active site and other regions of hMGL 37 .…”