Prospecting for cellulases from the marine environment: structure and function of novel GH7 and GH9 family glycoside hydrolases

Prospecting enzymes capable of digesting lignocellulosic biomass has focused on the relatively few types of microorganisms and animals that have wood-degrading capability. Of particular interest are the GH9-family enzymes that hydrolyze internal glycosidic bonds, and the GH7 cellobiohydrolase enzymes that processively hydrolyze cellulose polymers to cellobiose. We are exploring the rich resource of endogenous lignocellulose-degrading enzymes from the marine environment. Unlike animals such as termites that employ a complex community of microbial flora to produce digestive enzymes, the marine crustacean Limnoria quadripunctata has a sterile gut and produces all the necessary enzymatic machinery to efficiently digest these challenging substrates. A detailed characterization of Limnoria GH7 revealed remarkable properties such as highly acidic surface charge compared to the well-studied fungal GH7 from Hypocrea jecorina (Kern, et al, 2013, PNAS 110(25), 10189-94). In our latest studies, we have cloned, expressed and solved the X-ray structure of the GH7 from the water flea Daphnia pulex, one of the few other animal GH7s sequences available. This permits a structural and functional comparison of GH7 adaptations that have evolved in fresh water versus marine environments. Although GH7s have a very narrow distribution within the animal kingdom, GH9s are more widespread. In parallel work, we have characterized a GH9-family enzyme from the marine polychaete Perinereis brevicirris. A recent crystal structure reveals a similar acidic surface coat and we are now investigating the activity profile. From an industrial standpoint, robust, salt tolerant marine enzymes that efficiently degrade lignocellulosic substrates represent a new space for prospecting enhanced properties.