In Silico Analysis of Enzyme Surface and Glycosylation Effect as a Tool for Efficient Covalent Immobilisation of CalB and PGA on Sepabeads®

Abstract

This study presents a computational analysis of the structures of lipase B from Candida antarctica (CalB) and two penicillin G acylases (PGAs), from eukaryotic and prokaryotic sources, respectively. Molecular simulations were used to point out the regions of the enzymes that are prone to interact with immobilisation supports. In order to evaluate the accessibility of the active site, the location of the amino acid residues involved in the formation of covalent bonds with the polymers was visualised. The mapping of the distribution of hydrophobic and hydrophilic regions on the enzyme surface provided a view of the areas of the protein that can establish either hydrophobic or hydrophilic interactions with the carriers. Experimental data obtained from the immobilisation of the enzymes on supports bearing different chemical functionalities suggest the involvement of the glycan moiety in enzyme-polymer interactions. In the case of PGA the glycan moiety can constitute an extra site for the covalent linkage of the enzyme on the polymer.

Conformational Changes of Lipases in Aqueous Media: A Comparative Computational Study and Experimental Implications

“…The same concept applies for covalent immobilization on solid supports [15,16] and formulation of solid cross-linked aggregates (CLEA). [17] These immobilization methodologies generally imply the formation of multiple covalent bonds which confer higher rigidity to enzymes that are previously solubilized in aqueous buffer.…”

“…[15] In the first case (SPRIN immo P. cepacia1 and SPRIN immo CaLB1) the enzyme was immobilized in toluene [48] whereas in the second case the immobilization was performed in aqueous buffer. [15] Figure 11. Analysis of the geometry of the catalytic machinery of PcL, PaL and CrL.…”

“…Conformational changes were simulated in explicit water starting from their open active crystal structures. The study analyses changes occurring at three levels: (i) surface properties with special regards to variation of the exposure of the hydrophobic area in the proximity of the active site; [15] (ii) conformational changes of the lid domain and other domains possibly involved in the occlusion of the hydrophobic active site; (iii) conformational changes occurring at the catalytic machinery of the lipases. The objective was to observe different phenomena that might cause a decrease of activity of lipases when exposed to aqueous media.…”

Protein functionalized carbon nanomaterials for biomedical applications

“…In most cases, the choice of a suitable carrier is biased by particular polymers that have been shown to be effective with a large number of proteins [148]. Considering enzymes, this may result in non-optimal exploration of their catalytic efficiency.…”

Comparison of mesoporous silicate supports for the immobilisation and activity of cytochrome c and lipase

“…CALB is a hydrolase enzyme (33 kDa) [14] that is widely used in biocatalysis [15][16][17] due to its broad substrate range, high activity and stability. CALB has been successfully immobilized by several methods [18][19][20] onto various solid supports [21][22][23][24]. SBA-15 possesses a hexagonal structure with pore diameters of ca.…”