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Enhancement of Cellulose Degradation by Cattle Saliva.

Seki Y, Kikuchi Y, Kimura Y, Yoshimoto R, Takahashi M, Aburai K, Kanai Y, Ruike T, Iwabata K, Sugawara F, Sakai H, Abe M, Sakaguchi K - PLoS ONE (2015)

Bottom Line: Although cattle saliva, by itself, did not degrade cellulose, it enhanced the cellulase-catalyzed degradation of cellulose.Thus, the amount of reducing sugar produced increased approximately 2.9-fold by the addition of cattle saliva.We also found that non-enzymatic proteins, which were present in cattle saliva, were responsible for causing the enhancement effect.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Science and Technology, Tokyo University of Science, Noda, Chiba, Japan.

ABSTRACT
Saccharification of cellulose is a promising technique for producing alternative source of energy. However, the efficiency of conversion of cellulose into soluble sugar using any currently available methodology is too low for industrial application. Many additives, such as surfactants, have been shown to enhance the efficiency of cellulose-to-sugar conversion. In this study, we have examined first whether cattle saliva, as an additive, would enhance the cellulase-catalyzed hydrolysis of cellulose, and subsequently elucidated the mechanism by which cattle saliva enhanced this conversion. Although cattle saliva, by itself, did not degrade cellulose, it enhanced the cellulase-catalyzed degradation of cellulose. Thus, the amount of reducing sugar produced increased approximately 2.9-fold by the addition of cattle saliva. We also found that non-enzymatic proteins, which were present in cattle saliva, were responsible for causing the enhancement effect. Third, the mechanism of cattle saliva mediated enhancement of cellulase activity was probably similar to that of the canonical surfactants. Cattle saliva is available in large amounts easily and cheaply, and it can be used without further purification. Thus, cattle saliva could be a promising additive for efficient saccharification of cellulose on an industrial scale.

No MeSH data available.


Fractionation of cattle saliva proteins.(a) SDS-PAGE analysis of fractionated cattle saliva proteins. Proteins present in the cattle saliva were fractionated by gel filtration chromatography. An aliquot of each fraction was used for SDS-PAGE analysis, following which the acrylamide gel was stained with silver stain. Collected fractions were then divided as indicated (based on molecular weight distribution of proteins) into five sample groups (called here as sample A, B, C, D and E) for further use. (b) Enhancement effect of fractionated cattle saliva. Cellulase-catalyzed cellulose degradation assay (in triplicate) was performed using one of the fractionated saliva samples (A, B, C, D, or E) or Mix sample as an additive, and the amount of reducing sugar produced in each case was measured. Error bars indicate ± standard deviations. Values labeled with asterisk are statistically different as established by Student's t-test (P < 0.05).
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pone.0138902.g005: Fractionation of cattle saliva proteins.(a) SDS-PAGE analysis of fractionated cattle saliva proteins. Proteins present in the cattle saliva were fractionated by gel filtration chromatography. An aliquot of each fraction was used for SDS-PAGE analysis, following which the acrylamide gel was stained with silver stain. Collected fractions were then divided as indicated (based on molecular weight distribution of proteins) into five sample groups (called here as sample A, B, C, D and E) for further use. (b) Enhancement effect of fractionated cattle saliva. Cellulase-catalyzed cellulose degradation assay (in triplicate) was performed using one of the fractionated saliva samples (A, B, C, D, or E) or Mix sample as an additive, and the amount of reducing sugar produced in each case was measured. Error bars indicate ± standard deviations. Values labeled with asterisk are statistically different as established by Student's t-test (P < 0.05).

Mentions: Finally, to identify the protein responsible for the enhancement effect, we fractionated cattle saliva protein using gel-filtration chromatography. The fractionated proteins were subsequently analyzed by SDS-PAGE (Fig 5A). As can be seen, protein bands were present in fractions 9 through 20. Based on the molecular weight distribution of the fractionated proteins, we divided these factions into five sample groups, A through E, as follows: (A) fraction numbers 10 and 11 containing proteins whose molecular weights were more than 50 kDa; (B) fraction numbers 12 and 13 containing proteins with molecular weights of approximately 37 kDa; (C) fraction numbers 14 and 15 containing proteins with molecular weights of approximately 70, 25 and 15 kDa; (D) fraction numbers 16 and 17 containing proteins with molecular weights of approximately 25 and 15 kDa; and (E) fraction numbers 18 and 19 containing proteins whose molecular weights were similar to those in (D). We also prepared a mixed sample (Mix) by combining equal volumes of fractions A through E. We then used these samples in the cellulose degradation assay and the results are shown in Fig 5B. The amount of glucose produced using samples A, B and C as additives ranged from 0.140 mg/mL to 0.170 mg/mL, whereas the amount of glucose produced using samples D, E, Mix and Cellulase + Saliva (Saliva (+)) were approximately 0.190 mg/mL. In all cases, the amount of glucose produced using any one of these samples was higher than the amount of glucose produced using mixture containing only cellulase (Cellulose + Cellulase (Saliva (-))). Because all assay mixtures contained same amount of protein, the enhancement effect appeared to decrease slightly corresponding increase of the molecular weight of the protein.


Enhancement of Cellulose Degradation by Cattle Saliva.

Seki Y, Kikuchi Y, Kimura Y, Yoshimoto R, Takahashi M, Aburai K, Kanai Y, Ruike T, Iwabata K, Sugawara F, Sakai H, Abe M, Sakaguchi K - PLoS ONE (2015)

Fractionation of cattle saliva proteins.(a) SDS-PAGE analysis of fractionated cattle saliva proteins. Proteins present in the cattle saliva were fractionated by gel filtration chromatography. An aliquot of each fraction was used for SDS-PAGE analysis, following which the acrylamide gel was stained with silver stain. Collected fractions were then divided as indicated (based on molecular weight distribution of proteins) into five sample groups (called here as sample A, B, C, D and E) for further use. (b) Enhancement effect of fractionated cattle saliva. Cellulase-catalyzed cellulose degradation assay (in triplicate) was performed using one of the fractionated saliva samples (A, B, C, D, or E) or Mix sample as an additive, and the amount of reducing sugar produced in each case was measured. Error bars indicate ± standard deviations. Values labeled with asterisk are statistically different as established by Student's t-test (P < 0.05).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4581621&req=5

pone.0138902.g005: Fractionation of cattle saliva proteins.(a) SDS-PAGE analysis of fractionated cattle saliva proteins. Proteins present in the cattle saliva were fractionated by gel filtration chromatography. An aliquot of each fraction was used for SDS-PAGE analysis, following which the acrylamide gel was stained with silver stain. Collected fractions were then divided as indicated (based on molecular weight distribution of proteins) into five sample groups (called here as sample A, B, C, D and E) for further use. (b) Enhancement effect of fractionated cattle saliva. Cellulase-catalyzed cellulose degradation assay (in triplicate) was performed using one of the fractionated saliva samples (A, B, C, D, or E) or Mix sample as an additive, and the amount of reducing sugar produced in each case was measured. Error bars indicate ± standard deviations. Values labeled with asterisk are statistically different as established by Student's t-test (P < 0.05).
Mentions: Finally, to identify the protein responsible for the enhancement effect, we fractionated cattle saliva protein using gel-filtration chromatography. The fractionated proteins were subsequently analyzed by SDS-PAGE (Fig 5A). As can be seen, protein bands were present in fractions 9 through 20. Based on the molecular weight distribution of the fractionated proteins, we divided these factions into five sample groups, A through E, as follows: (A) fraction numbers 10 and 11 containing proteins whose molecular weights were more than 50 kDa; (B) fraction numbers 12 and 13 containing proteins with molecular weights of approximately 37 kDa; (C) fraction numbers 14 and 15 containing proteins with molecular weights of approximately 70, 25 and 15 kDa; (D) fraction numbers 16 and 17 containing proteins with molecular weights of approximately 25 and 15 kDa; and (E) fraction numbers 18 and 19 containing proteins whose molecular weights were similar to those in (D). We also prepared a mixed sample (Mix) by combining equal volumes of fractions A through E. We then used these samples in the cellulose degradation assay and the results are shown in Fig 5B. The amount of glucose produced using samples A, B and C as additives ranged from 0.140 mg/mL to 0.170 mg/mL, whereas the amount of glucose produced using samples D, E, Mix and Cellulase + Saliva (Saliva (+)) were approximately 0.190 mg/mL. In all cases, the amount of glucose produced using any one of these samples was higher than the amount of glucose produced using mixture containing only cellulase (Cellulose + Cellulase (Saliva (-))). Because all assay mixtures contained same amount of protein, the enhancement effect appeared to decrease slightly corresponding increase of the molecular weight of the protein.

Bottom Line: Although cattle saliva, by itself, did not degrade cellulose, it enhanced the cellulase-catalyzed degradation of cellulose.Thus, the amount of reducing sugar produced increased approximately 2.9-fold by the addition of cattle saliva.We also found that non-enzymatic proteins, which were present in cattle saliva, were responsible for causing the enhancement effect.

View Article: PubMed Central - PubMed

Affiliation: Research Institute for Science and Technology, Tokyo University of Science, Noda, Chiba, Japan.

ABSTRACT
Saccharification of cellulose is a promising technique for producing alternative source of energy. However, the efficiency of conversion of cellulose into soluble sugar using any currently available methodology is too low for industrial application. Many additives, such as surfactants, have been shown to enhance the efficiency of cellulose-to-sugar conversion. In this study, we have examined first whether cattle saliva, as an additive, would enhance the cellulase-catalyzed hydrolysis of cellulose, and subsequently elucidated the mechanism by which cattle saliva enhanced this conversion. Although cattle saliva, by itself, did not degrade cellulose, it enhanced the cellulase-catalyzed degradation of cellulose. Thus, the amount of reducing sugar produced increased approximately 2.9-fold by the addition of cattle saliva. We also found that non-enzymatic proteins, which were present in cattle saliva, were responsible for causing the enhancement effect. Third, the mechanism of cattle saliva mediated enhancement of cellulase activity was probably similar to that of the canonical surfactants. Cattle saliva is available in large amounts easily and cheaply, and it can be used without further purification. Thus, cattle saliva could be a promising additive for efficient saccharification of cellulose on an industrial scale.

No MeSH data available.