Journal Club:" Correlation of cellulase gene expression and cellulolytic acticity throughout the gut of the termite Reticulitermes flavipes."
It has been a while since the last contribution to this site. Today’s blog entry shall be a summary of a journal article that a couple of friends and I started recently to educate ourselves on topics related to bio-based ideas that are of interest to us. Recently, I came across a general article in the Scientist that talked about biofuels production. The article mentioned integrated biofuel productionpipelines that could include the breakdown of agricultural waste products containing cellulose.
Cellulose is a complex sugar, meaning that it consists of long-chains of individual simple sugar units such as glucose. The glucose that is released from the breakdown of cellulose could be used for bioethanol production. But it turns out that cellulose is a very sturdy material that is difficult and energy intensive to break down. One way to reduce the cost of break-down would be to rely on enzymes that are known as biological catalysts. Catalysts can reduce the activation energy which is the energy required to get a given reaction started. Most organisms do not have the necessary enzymes to break down cellulose. However, there are some organisms that do have them. Among these organisms are the termites.
Curious to learn more about cellulases, the enzymes that break the links between each glucose unit, in termites, we picked an article with the following title:
“Correlation of cellulase gene expression and cellulolytic acticity throughout the gut of the termite Reticulitermes flavipes.”
The authors in this paper used metagenomics approach to identify four open reading frames, predicted genes, in the sequences of the recently sequenced Reticulitermes flavipes. They then showed by PCR that these open reading frames are also expressed. The four genes are called Cell-1, Cell-2, Cell-3 and Cell-4. Cell-2 in particular was found to have at least one intron due to the fact that the genomic DNA sequence was larger than the cDNA sequence. Cell-1 was found to be highly similar to other termite cellulases. The other cellulose genes were more similar to symbiotic cellulases, meaning that these cellulases are not produced by termites themselves but by their prokaryotic symbiotic partners. Aligning Cell-2 to Cell-4, showed that only Cell-2 was lacking certain “tunnel forming loops” which are characteristic of exoglucanase. Cell-2 was then characterized as an endoglucanase.
The authors then looked at expression of the cellulose genes. They found that Cell-1 was mainly expressed in the fore-gut and salivary glands, while the remaining cellulases are expressed in the hind-gut. Characterizing endoglucanases, exoglucanases, and xylanases with respect to the location showed a strong bias of endoglucanases for foregut regions (~65% of all occurances) vs hindgut regions (~27%). For exoglucanases, the picture was reversed 61.5% occurance in hindgut and only ~14.0% in the foregut. Xylenases, on the other hand, were almost exclusively found in the hindgut. Endoglucanses break internal bonds to disrupt the crystalline structure of cellulose and expose individual cellulose polysaccharide chains. Exoglucanases cleave 2-4 units from the ends of the exposed chains produced by endocellulase, resulting tetrasaccharides or disaccharide such as cellobiose. Xylenases degrade beta-1,4-xylan into simple wood sugar. The distribution of Cell-1 through Cell-4 therefore mimics the logical breakdown of complex wood material into simple sugars.
Although most of the findings were not too surprising, I enjoyed reading this paper for the well-written introduction, and the logical approaches biologists can take to perform a relatively simple characterization of novel genes using simple bioinformatical tools. Performing these kinds of characterizations can help in the further optimization of wood degradation processes.
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Cellulose is a complex sugar, meaning that it consists of long-chains of individual simple sugar units such as glucose. The glucose that is released from the breakdown of cellulose could be used for bioethanol production. But it turns out that cellulose is a very sturdy material that is difficult and energy intensive to break down. One way to reduce the cost of break-down would be to rely on enzymes that are known as biological catalysts. Catalysts can reduce the activation energy which is the energy required to get a given reaction started. Most organisms do not have the necessary enzymes to break down cellulose. However, there are some organisms that do have them. Among these organisms are the termites.
Curious to learn more about cellulases, the enzymes that break the links between each glucose unit, in termites, we picked an article with the following title:
“Correlation of cellulase gene expression and cellulolytic acticity throughout the gut of the termite Reticulitermes flavipes.”
The authors in this paper used metagenomics approach to identify four open reading frames, predicted genes, in the sequences of the recently sequenced Reticulitermes flavipes. They then showed by PCR that these open reading frames are also expressed. The four genes are called Cell-1, Cell-2, Cell-3 and Cell-4. Cell-2 in particular was found to have at least one intron due to the fact that the genomic DNA sequence was larger than the cDNA sequence. Cell-1 was found to be highly similar to other termite cellulases. The other cellulose genes were more similar to symbiotic cellulases, meaning that these cellulases are not produced by termites themselves but by their prokaryotic symbiotic partners. Aligning Cell-2 to Cell-4, showed that only Cell-2 was lacking certain “tunnel forming loops” which are characteristic of exoglucanase. Cell-2 was then characterized as an endoglucanase.
The authors then looked at expression of the cellulose genes. They found that Cell-1 was mainly expressed in the fore-gut and salivary glands, while the remaining cellulases are expressed in the hind-gut. Characterizing endoglucanases, exoglucanases, and xylanases with respect to the location showed a strong bias of endoglucanases for foregut regions (~65% of all occurances) vs hindgut regions (~27%). For exoglucanases, the picture was reversed 61.5% occurance in hindgut and only ~14.0% in the foregut. Xylenases, on the other hand, were almost exclusively found in the hindgut. Endoglucanses break internal bonds to disrupt the crystalline structure of cellulose and expose individual cellulose polysaccharide chains. Exoglucanases cleave 2-4 units from the ends of the exposed chains produced by endocellulase, resulting tetrasaccharides or disaccharide such as cellobiose. Xylenases degrade beta-1,4-xylan into simple wood sugar. The distribution of Cell-1 through Cell-4 therefore mimics the logical breakdown of complex wood material into simple sugars.
Although most of the findings were not too surprising, I enjoyed reading this paper for the well-written introduction, and the logical approaches biologists can take to perform a relatively simple characterization of novel genes using simple bioinformatical tools. Performing these kinds of characterizations can help in the further optimization of wood degradation processes.
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