Abstract
Chitin is an abundant renewable polysaccharide, next only to cellulose. Chitinases are important for effective utilization of this biopolymer. Chitinase D from Serratia proteamaculans (SpChiD) is a single domain chitinase with both hydrolytic and transglycosylation (TG) activities. SpChiD had less of hydrolytic activity on insoluble polymeric chitin substrates due to the absence of auxiliary binding domains. We improved catalytic efficiency of SpChiD in degradation of insoluble chitin substrates by fusing with auxiliary domains like polycystic kidney disease (PKD) domain and chitin binding protein 21 (CBP21). Of the six different SpChiD fusion chimeras, two C-terminal fusions viz. ChiD+PKD and ChiD+CBP resulted in improved hydrolytic activity on α- and β-chitin, respectively. Time-course degradation of colloidal chitin also confirmed that these two C-terminal SpChiD fusion chimeras were more active than other chimeras. More TG products were produced for a longer duration by the fusion chimeras ChiD+PKD and PKD+ChiD+CBP.
Highlights
Chitin is the second most abundant natural polysaccharide consisting of (1–4) linked units of 2-acetamido-2-deoxy-β-D-glucopyranose (GlcNAc) in a linear form
E. coli Rosetta-gami II (DE3) cells harbouring the plasmids of SpChiD and the corresponding fusion chimeras were used for protein expression
We showed for the first time enhancement of hydrolytic activity on insoluble chitin polymer, for a single module glycosyl hydrolase family 18 (GH18) chitinase, with fusion of polycystic kidney disease (PKD) or chitin binding protein 21 (CBP21)
Summary
Chitin is the second most abundant natural polysaccharide consisting of (1–4) linked units of 2-acetamido-2-deoxy-β-D-glucopyranose (GlcNAc) in a linear form. It is insoluble in water and primarily exists in two crystalline (α- and β-) forms. The α-chitin contains sheets of tightly packed alternating parallel and antiparallel chains [1] and is abundant in the exoskeletons of arthropods, insects, fungi and yeast cell walls. The chains are arranged in parallel orientation in β-chitin [2], which occurs less frequently in nature and is often extracted from squid pens. Chemical hydrolysis of chitin results in CHOS with low degree of PLOS ONE | DOI:10.1371/journal.pone.0116823. Chemical hydrolysis of chitin results in CHOS with low degree of PLOS ONE | DOI:10.1371/journal.pone.0116823 January 23, 2015
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