Protein Crosslinking Methods

Gerrard, J.A. 2002. Protein-protein crosslinking in food Methods, consequences, applications. Trends Food Sci Technol 13(12) 389-397. 

Chitosan-based particles loaded with proteins can be obtained by both physical and chemical methods. However, the chemical crosslinking methods use toxic organic solvents (to make water/oil emulsions), heat, vigorous agitation or chemicals that might compromise the stability and biocompatibility of the proteins [93]. 

A technique used to obtain spherical particles is the spray-drying technique. The spray-drying method has reportedly provided small-size, protein-loaded chitosan-alginate microparticles with appropriate protein immobilization capacity [98]. It has been applied to BSA/sodium alginate/chitosan solutions to obtain microparticles [98]. Moreover, chitosan nanoparticles can be obtained by ionic crosslinking method. It has produced nanoparticles based on ionic gelation process of tripolyphosphate (TPP) and chitosan [99]. Nevertheless, another physical method called layer-by-layer can be used to form chitosan particles. Through the layer-by-layer technique submicron (40 nm) capsules with poly(L-aspartic acid) and chitosan for transmucosal delivery of proteins and peptides have been obtained [100]. 

Numerous organisms, both marine and terrestrial, produce protein toxins. These are typically relatively small, and rich in disulfide crosslinks. Since they are often difficult to crystallize, relatively few structures from this class of proteins are known. In the past five years two dimensional NMR methods have developed to the point where they can be used to determine the solution structures of small proteins and nucleic acids. We have analyzed the structures of toxins II and III of RadiarUhus paumotensis using this approach. We find that the dominant structure is )9-sheet, with the strands connected by loops of irregular structure. Most of the residues which have been determined to be important for toxicity are contained in one of the loops. The general methods used for structure analysis will be described, and the structures of the toxins RpII and RpIII will be discussed and compared with homologous toxins from other anemone species. 

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