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Affinity maturation generates greatly improved xyloglucan-specific carbohydrate binding modules

von Schantz, Laura LU ; Gullfot, Fredrika; Scheer, Sebastian; Filonova, Lada; Cicortas Gunnarsson, Lavinia LU ; Flint, James E; Daniel, Geoffrey; Nordberg Karlsson, Eva LU ; Brumer, Harry and Ohlin, Mats LU (2009) In BMC Biotechnology 9.
Abstract
BACKGROUND: Molecular evolution of carbohydrate binding modules (CBM) is a new approach for the generation of glycan-specific molecular probes. To date, the possibility of performing affinity maturation on CBM has not been investigated. In this study we show that binding characteristics such as affinity can be improved for CBM generated from the CBM4-2 scaffold by using random mutagenesis in combination with phage display technology. RESULTS: Two modified proteins with greatly improved affinity for xyloglucan, a key polysaccharide abundant in the plant kingdom crucial for providing plant support, were generated. Both improved modules differ from other existing xyloglucan probes by binding to galactose-decorated subunits of xyloglucan. The... (More)
BACKGROUND: Molecular evolution of carbohydrate binding modules (CBM) is a new approach for the generation of glycan-specific molecular probes. To date, the possibility of performing affinity maturation on CBM has not been investigated. In this study we show that binding characteristics such as affinity can be improved for CBM generated from the CBM4-2 scaffold by using random mutagenesis in combination with phage display technology. RESULTS: Two modified proteins with greatly improved affinity for xyloglucan, a key polysaccharide abundant in the plant kingdom crucial for providing plant support, were generated. Both improved modules differ from other existing xyloglucan probes by binding to galactose-decorated subunits of xyloglucan. The usefulness of the evolved binders was verified by staining of plant sections, where they performed better than the xyloglucan-binding module from which they had been derived. They discriminated non-fucosylated from fucosylated xyloglucan as shown by their ability to stain only the endosperm, rich in non-fucosylated xyloglucan, but not the integument rich in fucosylated xyloglucan, on tamarind seed sections. CONCLUSION: We conclude that affinity maturation of CBM selected from molecular libraries based on the CBM4-2 scaffold is possible and has the potential to generate new analytical tools for detection of plant carbohydrates. (Less)
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author
organization
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type
Contribution to journal
publication status
published
subject
in
BMC Biotechnology
volume
9
publisher
BioMed Central
external identifiers
  • wos:000272337000001
  • scopus:71049184894
ISSN
1472-6750
DOI
10.1186/1472-6750-9-92
language
English
LU publication?
yes
id
b11d150b-8627-4453-8451-355c8cc43e59 (old id 1487461)
date added to LUP
2009-10-12 13:24:35
date last changed
2017-09-03 04:01:56
@article{b11d150b-8627-4453-8451-355c8cc43e59,
  abstract     = {BACKGROUND: Molecular evolution of carbohydrate binding modules (CBM) is a new approach for the generation of glycan-specific molecular probes. To date, the possibility of performing affinity maturation on CBM has not been investigated. In this study we show that binding characteristics such as affinity can be improved for CBM generated from the CBM4-2 scaffold by using random mutagenesis in combination with phage display technology. RESULTS: Two modified proteins with greatly improved affinity for xyloglucan, a key polysaccharide abundant in the plant kingdom crucial for providing plant support, were generated. Both improved modules differ from other existing xyloglucan probes by binding to galactose-decorated subunits of xyloglucan. The usefulness of the evolved binders was verified by staining of plant sections, where they performed better than the xyloglucan-binding module from which they had been derived. They discriminated non-fucosylated from fucosylated xyloglucan as shown by their ability to stain only the endosperm, rich in non-fucosylated xyloglucan, but not the integument rich in fucosylated xyloglucan, on tamarind seed sections. CONCLUSION: We conclude that affinity maturation of CBM selected from molecular libraries based on the CBM4-2 scaffold is possible and has the potential to generate new analytical tools for detection of plant carbohydrates.},
  articleno    = {92},
  author       = {von Schantz, Laura and Gullfot, Fredrika and Scheer, Sebastian and Filonova, Lada and Cicortas Gunnarsson, Lavinia and Flint, James E and Daniel, Geoffrey and Nordberg Karlsson, Eva and Brumer, Harry and Ohlin, Mats},
  issn         = {1472-6750},
  language     = {eng},
  publisher    = {BioMed Central},
  series       = {BMC Biotechnology},
  title        = {Affinity maturation generates greatly improved xyloglucan-specific carbohydrate binding modules},
  url          = {http://dx.doi.org/10.1186/1472-6750-9-92},
  volume       = {9},
  year         = {2009},
}