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Directed evolution of Aspergillus niger glucoamylase to increase thermostability.

McDaniel A, Fuchs E, Liu Y, Ford C - Microb Biotechnol (2008)

Bottom Line: Irreversible inactivation tests indicated that CR2-1 increases the free energy of thermoinactivation at 80°C by 10 kJ mol(-1) compared with that of wild-type GA.Thus, CR2-1 is more thermostable (by 5 kJ mol(-1) at 80°C) than the most thermostable A. niger GA variant previously described, THS8.In addition, Val59Ala and Glu408Lys were shown to individually increase the thermostability in GA variants by 1 and 2 kJ mol(-1), respectively, at 80°C.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science and Human Nutrition, Iowa State University, Ames, IA 50011, USA.

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Mentions: Site‐directed mutagenesis was used to recreate Val88Ile, Ser211Pro, Asp293Ala, Tyr402Phe and Glu408Lys singly in an otherwise wild‐type GA background for further thermostability tests. Thr390Ser was not isolated in a wild‐type GA background for further thermostability testing due to the similarity of the replacement amino acid side‐group, and its likely neutrality with respect to thermostability. A thermostability plate assay was used to test the individual phenotypic effect of the single mutations Val88Ile, Ser211Pro, Asp293Ala, Tyr402Phe and Glu408Lys compared with wild‐type GA, Ser30Pro (Allen et al., 1998), RE15 and SRE15 (Fig. 3). Of the new single mutations, only Glu408Lys showed a slightly increased resistance to thermoinactivation compared with wild‐type GA. In contrast, both RE15 and SRE15 showed substantially increased thermostability.


Directed evolution of Aspergillus niger glucoamylase to increase thermostability.

McDaniel A, Fuchs E, Liu Y, Ford C - Microb Biotechnol (2008)

© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3815294&req=5

Mentions: Site‐directed mutagenesis was used to recreate Val88Ile, Ser211Pro, Asp293Ala, Tyr402Phe and Glu408Lys singly in an otherwise wild‐type GA background for further thermostability tests. Thr390Ser was not isolated in a wild‐type GA background for further thermostability testing due to the similarity of the replacement amino acid side‐group, and its likely neutrality with respect to thermostability. A thermostability plate assay was used to test the individual phenotypic effect of the single mutations Val88Ile, Ser211Pro, Asp293Ala, Tyr402Phe and Glu408Lys compared with wild‐type GA, Ser30Pro (Allen et al., 1998), RE15 and SRE15 (Fig. 3). Of the new single mutations, only Glu408Lys showed a slightly increased resistance to thermoinactivation compared with wild‐type GA. In contrast, both RE15 and SRE15 showed substantially increased thermostability.

Bottom Line: Irreversible inactivation tests indicated that CR2-1 increases the free energy of thermoinactivation at 80°C by 10 kJ mol(-1) compared with that of wild-type GA.Thus, CR2-1 is more thermostable (by 5 kJ mol(-1) at 80°C) than the most thermostable A. niger GA variant previously described, THS8.In addition, Val59Ala and Glu408Lys were shown to individually increase the thermostability in GA variants by 1 and 2 kJ mol(-1), respectively, at 80°C.

View Article: PubMed Central - PubMed

Affiliation: Department of Food Science and Human Nutrition, Iowa State University, Ames, IA 50011, USA.

Show MeSH
Related in: MedlinePlus