Abstract
BackgroundAlthough many hyperthermophilic endoglucanases have been reported from archaea and bacteria, a complete survey and classification of all sequences in these species from disparate evolutionary groups, and the relationship between their molecular structures and functions are lacking. The completion of several high-quality gene or genome sequencing projects provided us with the unique opportunity to make a complete assessment and thorough comparative analysis of the hyperthermophilic endoglucanases encoded in archaea and bacteria.ResultsStructure alignment of the 19 hyperthermophilic endoglucanases from archaea and bacteria which grow above 80°C revealed that Gly30, Pro63, Pro83, Trp115, Glu131, Met133, Trp135, Trp175, Gly227 and Glu229 are conserved amino acid residues. In addition, the average percentage composition of residues cysteine and histidine of 19 endoglucanases is only 0.28 and 0.74 while it is high in thermophilic or mesophilic one. It can be inferred from the nodes that there is a close relationship among the 19 protein from hyperthermophilic bacteria and archaea based on phylogenetic analysis. Among these conserved amino acid residues, as far as Cel12B concerned, two Glu residues might be the catalytic nucleophile and proton donor, Gly30, Pro63, Pro83 and Gly227 residues might be necessary to the thermostability of protein, and Trp115, Met133, Trp135, Trp175 residues is related to the binding of substrate. Site-directed mutagenesis results reveal that Pro63 and Pro83 contribute to the thermostability of Cel12B and Met133 is confirmed to have role in enhancing the binding of substrate.ConclusionsThe conserved acids have been shown great importance to maintain the structure, thermostability, as well as the similarity of the enzymatic properties of those proteins. We have made clear the function of these conserved amino acid residues in Cel12B protein, which is helpful in analyzing other undetailed molecular structure and transforming them with site directed mutagenesis, as well as providing the theoretical basis for degrading cellulose from woody and herbaceous plants.
Highlights
Many hyperthermophilic endoglucanases have been reported from archaea and bacteria, a complete survey and classification of all sequences in these species from disparate evolutionary groups, and the relationship between their molecular structures and functions are lacking
We identified and classified in this study a higher number of hyperthermophilic endoglucanase amino acids from the GHF12 than previously reported, allowing us to identify their relationships based on the phylogenetic clustering
We searched endoglucanase sequences in several plants, bacteria, fungi and algae sequences including the sequences of the R. speratus, using the protein BLAST search engine with a variety of endoglucanase amino acid sequences as queries for most of the thermophilic endoglucanase, else using endoglucanase as a keyword for searching other amino acid sequences of endoglucanase (Table 1)
Summary
Many hyperthermophilic endoglucanases have been reported from archaea and bacteria, a complete survey and classification of all sequences in these species from disparate evolutionary groups, and the relationship between their molecular structures and functions are lacking. The completion of several high-quality gene or genome sequencing projects provided us with the unique opportunity to make a complete assessment and thorough comparative analysis of the hyperthermophilic endoglucanases encoded in archaea and bacteria. Based on amino acid sequence homologies and hydrophobic cluster analysis, hyperthermophilic endoglucanases obtained from extremophiles, which are widely distributed in terrestrial and marine hydrothermal areas, as well as in deep subsurface oil reservoirs, have been classified into GHF12 [7,8,9,10,11,12,13,14]. Many hyperthermophilic endoglucanases gene which have been cloned were found in some heat-tolerant bacteria [16]. Those hyperthermophilic endoglucanases have a common feature that the amino acid sequences are mostly relatively short (less than 400 amino acid residues)
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