Abstract
ABSTRACT : Anaerobic rumen fungi have been regarded as good genetic resources for enzyme production which might be useful for feed supplements, bio-energy production, bio-remediation and other industrial purposes. In this study, an expressed sequence tag (EST) library of the rumen anaerobic fungus Neocallimastix frontalis was constructed and functional genes from the EST library were analyzed to elucidate carbohydrate metabolism of anaerobic fungi. From 10,080 acquired clones, 9,569 clones with average size of 628 bp were selected for analysis. After the assembling process, 1,410 contigs were assembled and 1,369 sequences remained as singletons. 1,192 sequences were matched with proteins in the public data base with known function and 693 of them were matched with proteins isolated from fungi. One hundred and fifty four sequences were classified as genes related with biological process and 328 sequences were classified as genes related with cellular components. Most of the enzymes in the pathway of glucose metabolism were successfully isolated via construction of 10,080 ESTs. Four kinds of hemi-cellulase were isolated such as mannanase, xylose isomerase, xylan esterase, and xylanase. Five β-glucosidases with at least three different conserved domain structures were isolated. Ten cellulases with at least five different conserved domain structures were isolated. This is the first solid data supporting the expression of a multiple enzyme system in the fungus
Highlights
Ruminant animals use fibrous plant materials as feed by degrading insoluble polysaccharides during microbial fermentation
The genera Neocallimastix, Orpinomyces and Piromyces are members of Family Neocallimastigaceae and they are very close in genetic relation and the genes encoding glucose hydrolyzing enzymes are highly homologous among anaerobic fungi (Harhangi et al, 2003)
There might be a strong possibility that N. frontalis secrete more than 4 glucose hydrolyzing enzymes which have not been detected
Summary
Ruminant animals use fibrous plant materials as feed by degrading insoluble polysaccharides during microbial fermentation. The genera Neocallimastix, Orpinomyces and Piromyces are members of Family Neocallimastigaceae and they are very close in genetic relation and the genes encoding glucose hydrolyzing enzymes are highly homologous among anaerobic fungi (Harhangi et al, 2003). These genes are even homologous to those in anaerobic bacteria due to horizontal gene transfer (Garcia-Vallve et al, 2000). The purpose of this study was to isolate functional genes related with carbohydrate metabolism from the anaerobic rumen fungus N. frontalis using high throughput EST analysis for further research
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