Abstract
The glycoside hydrolase 9 superfamily, mainly comprising the endoglucanases, is represented in all three domains of life. The current division of GH9 enzymes, into three subclasses, namely A, B, and C, is centered on parameters derived from sequence information alone. However, this classification is ambiguous, and is limited by the paralogous ancestry of classes B and C endoglucanases, and paucity of biochemical and structural data. Here, we extend this classification schema to putative GH9 endoglucanases present in green plants, with an emphasis on identifying novel members of the class C subset. These enzymes cleave the β(1 → 4) linkage between non-terminal adjacent D-glucopyranose residues, in both, amorphous and crystalline regions of cellulose. We utilized non redundant plant GH9 enzymes with characterized molecular data, as the training set to construct Hidden Markov Models (HMMs) and train an Artificial Neural Network (ANN). The parameters that were used for predicting dominant enzyme function, were derived from this training set, and subsequently refined on 147 sequences with available expression data. Our knowledge-based approach, can ascribe differential endoglucanase activity (A, B, or C) to a query sequence with high confidence, and was used to construct a local repository of class C GH9 endoglucanases (GH9C = 241) from 32 sequenced green plants.
Highlights
Cellulose, a straight chain organic polymer of several hundreds of repeating disaccharide units of D-glucopyranose in a β (1 → 4) glycosidic linkage, is present in the primary cell wall of plants, algae, and oomycetes, and is a critical component of bacterial biofilms (Updegraff, 1969; Yoshida Y. et al, 2006; Reardon-Robinson et al, 2014; Augimeri et al, 2015)
Our objective was to scan this data for highly probable class C sequences, a mathematical filter was developed to screen these on the basis of the Hidden Markov Model (HMM) scores of the included profiles
The phylogenetic clades were similar for both sets of profiles (Figures 3A,B), profile pairs were averaged: HMMGH9A : = average (HMMGH9A1D, HMMGH9A3D) (Def.13)
Summary
A straight chain organic polymer of several hundreds of repeating disaccharide units of D-glucopyranose in a β (1 → 4) glycosidic linkage, is present in the primary cell wall of plants, algae, and oomycetes, and is a critical component of bacterial biofilms (Updegraff, 1969; Yoshida Y. et al, 2006; Reardon-Robinson et al, 2014; Augimeri et al, 2015). The. Identification of Class C GH9 Endoglucanases in Plants cohesive structure of cellulose is aided, by a rich network of non-covalent hydrogen bonds between the hydroxyl (−OH−) groups of the glucose moieties of its constituent microfibrils. The presence of cellulose in primary cell walls, whilst protective and strength conferring, is important in the development and maintenance of bacterial biofilms for host interaction (Rhizobiaceae, Enterobacteriaceae, Acetobacteriaceae, etc.; Augimeri et al, 2015). This stability of cellulose, mandates a breakdown into constituent mono- and oligo-saccharides, prior to major patho-physiological events in plants. Plant development, along with stress adaptor mechanisms, are critically dependent on the digestion of cellulose (del Campillo et al, 2012; Kundu, 2015a)
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