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How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity

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How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity. / Cuskin, Fiona; Flint, James E.; Gloster, Tracey M.; Morland, Carl; Basle, Arnaud; Henrissat, Bernard; Coutinho, Pedro M.; Strazzulli, Andrea; Solovyova, Alexandra S.; Davies, Gideon J.; Gilbert, Harry J.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 109, No. 51, 18.12.2012, p. 20889-20894.

Research output: Contribution to journalArticlepeer-review

Harvard

Cuskin, F, Flint, JE, Gloster, TM, Morland, C, Basle, A, Henrissat, B, Coutinho, PM, Strazzulli, A, Solovyova, AS, Davies, GJ & Gilbert, HJ 2012, 'How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity', Proceedings of the National Academy of Sciences of the United States of America, vol. 109, no. 51, pp. 20889-20894. https://doi.org/10.1073/pnas.1212034109

APA

Cuskin, F., Flint, J. E., Gloster, T. M., Morland, C., Basle, A., Henrissat, B., Coutinho, P. M., Strazzulli, A., Solovyova, A. S., Davies, G. J., & Gilbert, H. J. (2012). How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity. Proceedings of the National Academy of Sciences of the United States of America, 109(51), 20889-20894. https://doi.org/10.1073/pnas.1212034109

Vancouver

Cuskin F, Flint JE, Gloster TM, Morland C, Basle A, Henrissat B et al. How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity. Proceedings of the National Academy of Sciences of the United States of America. 2012 Dec 18;109(51):20889-20894. https://doi.org/10.1073/pnas.1212034109

Author

Cuskin, Fiona ; Flint, James E. ; Gloster, Tracey M. ; Morland, Carl ; Basle, Arnaud ; Henrissat, Bernard ; Coutinho, Pedro M. ; Strazzulli, Andrea ; Solovyova, Alexandra S. ; Davies, Gideon J. ; Gilbert, Harry J. / How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity. In: Proceedings of the National Academy of Sciences of the United States of America. 2012 ; Vol. 109, No. 51. pp. 20889-20894.

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@article{2653bdc29be140bab5078070296fe665,
title = "How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity",
abstract = "Noncatalytic carbohydrate binding modules (CBMs) are components of glycoside hydrolases that attack generally inaccessible substrates. CBMs mediate a two-to fivefold elevation in the activity of endo-acting enzymes, likely through increasing the concentration of the appended enzymes in the vicinity of the substrate. The function of CBMs appended to exo-acting glycoside hydrolases is unclear because their typical endo-binding mode would not fulfill a targeting role. Here we show that the Bacillus subtilis exo-acting beta-fructosidase SacC, which specifically hydrolyses levan, contains the founding member of CBM family 66 (CBM66). The SacC-derived CBM66 (BsCBM66) targets the terminal fructosides of the major fructans found in nature. The crystal structure of BsCBM66 in complex with ligands reveals extensive interactions with the terminal fructose moiety (Fru-3) of levantriose but only limited hydrophobic contacts with Fru-2, explaining why the CBM displays broad specificity. Removal of BsCBM66 from SacC results in a similar to 100-fold reduction in activity against levan. The truncated enzyme functions as a nonspecific beta-fructosidase displaying similar activity against beta-2,1- and beta-2,6-linked fructans and their respective fructooligosaccharides. Conversely, appending BsCBM66 to BT3082, a nonspecific beta-fructosidase from Bacteroides thetaiotaomicron, confers exolevanase activity on the enzyme. We propose that BsCBM66 confers specificity for levan, a branched fructan, through an {"}avidity{"} mechanism in which the CBM and the catalytic module target the termini of different branches of the same polysaccharide molecule. This report identifies a unique mechanism by which CBMs modulate enzyme function, and shows how specificity can be tailored by integrating nonspecific catalytic and binding modules into a single enzyme.",
keywords = "BACILLUS-SUBTILIS, PURIFICATION, biofuels, CELLOBIOHYDROLASES, lectins, X-ray crystallography, FAMILIES, prebiotics, CELLULOSE DEGRADATION, RECOGNITION, INVERTASE, DOMAIN, LECTINS, isothermal titration calorimetry, DECONSTRUCTION",
author = "Fiona Cuskin and Flint, {James E.} and Gloster, {Tracey M.} and Carl Morland and Arnaud Basle and Bernard Henrissat and Coutinho, {Pedro M.} and Andrea Strazzulli and Solovyova, {Alexandra S.} and Davies, {Gideon J.} and Gilbert, {Harry J.}",
year = "2012",
month = dec,
day = "18",
doi = "10.1073/pnas.1212034109",
language = "English",
volume = "109",
pages = "20889--20894",
journal = "Proceedings of the National Academy of Sciences of the United States of America",
issn = "0027-8424",
publisher = "National Academy of Sciences",
number = "51",

}

RIS (suitable for import to EndNote) - Download

TY - JOUR

T1 - How nature can exploit nonspecific catalytic and carbohydrate binding modules to create enzymatic specificity

AU - Cuskin, Fiona

AU - Flint, James E.

AU - Gloster, Tracey M.

AU - Morland, Carl

AU - Basle, Arnaud

AU - Henrissat, Bernard

AU - Coutinho, Pedro M.

AU - Strazzulli, Andrea

AU - Solovyova, Alexandra S.

AU - Davies, Gideon J.

AU - Gilbert, Harry J.

PY - 2012/12/18

Y1 - 2012/12/18

N2 - Noncatalytic carbohydrate binding modules (CBMs) are components of glycoside hydrolases that attack generally inaccessible substrates. CBMs mediate a two-to fivefold elevation in the activity of endo-acting enzymes, likely through increasing the concentration of the appended enzymes in the vicinity of the substrate. The function of CBMs appended to exo-acting glycoside hydrolases is unclear because their typical endo-binding mode would not fulfill a targeting role. Here we show that the Bacillus subtilis exo-acting beta-fructosidase SacC, which specifically hydrolyses levan, contains the founding member of CBM family 66 (CBM66). The SacC-derived CBM66 (BsCBM66) targets the terminal fructosides of the major fructans found in nature. The crystal structure of BsCBM66 in complex with ligands reveals extensive interactions with the terminal fructose moiety (Fru-3) of levantriose but only limited hydrophobic contacts with Fru-2, explaining why the CBM displays broad specificity. Removal of BsCBM66 from SacC results in a similar to 100-fold reduction in activity against levan. The truncated enzyme functions as a nonspecific beta-fructosidase displaying similar activity against beta-2,1- and beta-2,6-linked fructans and their respective fructooligosaccharides. Conversely, appending BsCBM66 to BT3082, a nonspecific beta-fructosidase from Bacteroides thetaiotaomicron, confers exolevanase activity on the enzyme. We propose that BsCBM66 confers specificity for levan, a branched fructan, through an "avidity" mechanism in which the CBM and the catalytic module target the termini of different branches of the same polysaccharide molecule. This report identifies a unique mechanism by which CBMs modulate enzyme function, and shows how specificity can be tailored by integrating nonspecific catalytic and binding modules into a single enzyme.

AB - Noncatalytic carbohydrate binding modules (CBMs) are components of glycoside hydrolases that attack generally inaccessible substrates. CBMs mediate a two-to fivefold elevation in the activity of endo-acting enzymes, likely through increasing the concentration of the appended enzymes in the vicinity of the substrate. The function of CBMs appended to exo-acting glycoside hydrolases is unclear because their typical endo-binding mode would not fulfill a targeting role. Here we show that the Bacillus subtilis exo-acting beta-fructosidase SacC, which specifically hydrolyses levan, contains the founding member of CBM family 66 (CBM66). The SacC-derived CBM66 (BsCBM66) targets the terminal fructosides of the major fructans found in nature. The crystal structure of BsCBM66 in complex with ligands reveals extensive interactions with the terminal fructose moiety (Fru-3) of levantriose but only limited hydrophobic contacts with Fru-2, explaining why the CBM displays broad specificity. Removal of BsCBM66 from SacC results in a similar to 100-fold reduction in activity against levan. The truncated enzyme functions as a nonspecific beta-fructosidase displaying similar activity against beta-2,1- and beta-2,6-linked fructans and their respective fructooligosaccharides. Conversely, appending BsCBM66 to BT3082, a nonspecific beta-fructosidase from Bacteroides thetaiotaomicron, confers exolevanase activity on the enzyme. We propose that BsCBM66 confers specificity for levan, a branched fructan, through an "avidity" mechanism in which the CBM and the catalytic module target the termini of different branches of the same polysaccharide molecule. This report identifies a unique mechanism by which CBMs modulate enzyme function, and shows how specificity can be tailored by integrating nonspecific catalytic and binding modules into a single enzyme.

KW - BACILLUS-SUBTILIS

KW - PURIFICATION

KW - biofuels

KW - CELLOBIOHYDROLASES

KW - lectins

KW - X-ray crystallography

KW - FAMILIES

KW - prebiotics

KW - CELLULOSE DEGRADATION

KW - RECOGNITION

KW - INVERTASE

KW - DOMAIN

KW - LECTINS

KW - isothermal titration calorimetry

KW - DECONSTRUCTION

UR - http://www.scopus.com/inward/record.url?scp=84871379886&partnerID=8YFLogxK

U2 - 10.1073/pnas.1212034109

DO - 10.1073/pnas.1212034109

M3 - Article

VL - 109

SP - 20889

EP - 20894

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

SN - 0027-8424

IS - 51

ER -