Journal ArticleDOI
Structural origins of the interfacial activation in Thermomyces (Humicola) lanuginosa lipase.
Andrzej M. Brzozowski,H Savage,Chandra S. Verma,Johan P. Turkenburg,D M Lawson,Allan Svendsen,Shamkant Anant Patkar +6 more
TLDR
Five new Thermomyces (formerly Humicola) lanuginosa lipase (TlL) crystal structures have been solved and compared with four previously reported structures of this enzyme, suggesting that the sequence of the structural changes, as exemplified in various TlL crystal structures, mirror those that may occur during interfacial activation.Abstract:
The already known X-ray structures of lipases provide little evidence about initial, discrete structural steps occurring in the first phases of their activation in the presence of lipids (process referred to as interfacial activation). To address this problem, five new Thermomyces (formerly Humicola) lanuginosa lipase (TlL) crystal structures have been solved and compared with four previously reported structures of this enzyme. The bias coming from different crystallization media has been minimized by the growth of all crystals under the same crystallization conditions, in the presence of detergent/lipid analogues, with low or high ionic strength as the only main variable. Resulting structures and their characteristic features allowed the identification of three structurally distinct species of this enzyme: low activity form (LA), activated form (A), and fully Active (FA) form. The isomerization of the Cys268-Cys22 disulfide, synchronized with the formation of a new, short alpha(0) helix and flipping of the Arg84 (Arginine switch) located in the lid's proximal hinge, have been postulated as the key, structural factors of the initial transitions between LA and A forms. The experimental results were supplemented by theoretical calculations. The magnitude of the activation barrier between LA (ground state) and A (end state) forms of TlL (10.6 kcal/mol) is comparable to the enthalpic barriers typical for ring flips and disulfide isomerizations at ambient temperatures. This suggests that the sequence of the structural changes, as exemplified in various TlL crystal structures, mirror those that may occur during interfacial activation.read more
Citations
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Journal ArticleDOI
Modifying enzyme activity and selectivity by immobilization.
Rafael C. Rodrigues,Claudia Ortiz,Ángel Berenguer-Murcia,Rodrigo Torres,Roberto Fernandez-Lafuente +4 more
TL;DR: In this tutorial review, some of the main reasons that may produce an improvement in enzyme activity, specificity or selectivity, either real or apparent, due to immobilization are listed.
Journal ArticleDOI
Interfacial Enzymology: The Secreted Phospholipase A2-Paradigm
Journal ArticleDOI
The Lid Domain in Lipases: Structural and Functional Determinant of Enzymatic Properties
TL;DR: Improved computational models can now rationalize observations by relating it to the mobility of the lid domain and summarized and critically evaluated the most recent developments in experimental and computational research on lipase lids.
Journal ArticleDOI
Versatility of glutaraldehyde to immobilize lipases: Effect of the immobilization protocol on the properties of lipase B from Candida antarctica
TL;DR: 5 different CALB biocatalysts were prepared following the previous described protocols, and its stability and activity, pH/activity profile and specificity versus R and S methyl mandelate were analyzed, suggesting that the immobilization protocol may greatly affect the final effect of a chemical modification on the enzyme properties.
Journal ArticleDOI
Identification and Characterization of Bacterial Cutinase
TL;DR: The cutinase from Thermobifida fusca was induced by cutin and purified to homogeneity by following p-nitrophenyl butyrate hydrolyzing activity, the first report of cut inase encoding genes from bacterial sources.
References
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Journal ArticleDOI
CHARMM: A program for macromolecular energy, minimization, and dynamics calculations
Bernard R. Brooks,Robert E. Bruccoleri,Barry D. Olafson,David J. States,S. Swaminathan,Martin Karplus +5 more
TL;DR: The CHARMM (Chemistry at Harvard Macromolecular Mechanics) as discussed by the authors is a computer program that uses empirical energy functions to model macromolescular systems, and it can read or model build structures, energy minimize them by first- or second-derivative techniques, perform a normal mode or molecular dynamics simulation, and analyze the structural, equilibrium, and dynamic properties determined in these calculations.
Journal ArticleDOI
Refinement of macromolecular structures by the maximum-likelihood method.
TL;DR: The likelihood function for macromolecular structures is extended to include prior phase information and experimental standard uncertainties and the results derived are consistently better than those obtained from least-squares refinement.
Journal ArticleDOI
AMoRe: an automated package for molecular replacement
TL;DR: In this paper, a new molecular-replacement package is presented, which is an improvement on conventional methods, based on more powerful algorithms and a new conception that enables automation and rapid solution.
Journal ArticleDOI
MOLREP: an Automated Program for Molecular Replacement
Alexei A. Vagin,Alexei Teplyakov +1 more
TL;DR: MOLREP as mentioned in this paper is an automated program for molecular replacement which utilizes effective new approaches in data processing and rotational and translational searching, such as automatic choice of all parameters, scaling by Patterson origin peaks and soft resolution cut-off.
Journal ArticleDOI
Conjugate peak refinement: an algorithm for finding reaction paths and accurate transition states in systems with many degrees of freedom
Stefan Fischer,Martin Karplus +1 more
TL;DR: In this paper, an algorithm is presented for determining multi-dimensional reaction coordinates between two known conformers, which is suitable for the study of complex isomerization reactions, including allosteric transitions in proteins and more general conformational changes of macromolecules.