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Engineering protein-carbohydrate interactions - lessons from natural and evolved carbohydrate binding modules

von Schantz, Laura LU (2012)
Abstract
Protein-carbohydrate interactions are of central role in all living organism. In the studies presented in this thesis several strategies for engineering such interactions have been investigated. In the first study the binding affinity of a carbohydrate binding module (CBM) was improved a 100-fold through affinity maturation. Also, in that study it was discovered that all the selected, matured mutants carried a mutation in common (glutamic acid), which was shown to be responsible for the increased affinity.

Furthermore, the binding interaction of two different CBM was analyzed with x-ray crystallography. One of the CBM can only bind xylans, while the other binds also beta-glucans and xyloglucan. The crystal structures revealed that... (More)
Protein-carbohydrate interactions are of central role in all living organism. In the studies presented in this thesis several strategies for engineering such interactions have been investigated. In the first study the binding affinity of a carbohydrate binding module (CBM) was improved a 100-fold through affinity maturation. Also, in that study it was discovered that all the selected, matured mutants carried a mutation in common (glutamic acid), which was shown to be responsible for the increased affinity.

Furthermore, the binding interaction of two different CBM was analyzed with x-ray crystallography. One of the CBM can only bind xylans, while the other binds also beta-glucans and xyloglucan. The crystal structures revealed that the binding cleft of the multi-specific CBM is flexible, permitting reorientation of side-chains to avoid steric clashes. Also the multi-specific CBM harbored an important phenylalanine that due to its chemical composition has a pi-electron surface and can interact with hydrogen atoms on the different ligands it recognizes. The specific CBM on the other hand harbor a leucine in this position and can only interact with xylans.

In the last study, the capacity of CBM for use as analytical tools in the characterization of the distribution pattern of cellulose derivatives was investigated. The study showed that some CBM are able to distinguish between cellulose derivatives with different substitution distribution. This is a potentially new application for CBM.

In all, the thesis demonstrates strategies in which carbohydrate binding proteins can be generated, improved and utilized. (Less)
Please use this url to cite or link to this publication:
author
supervisor
opponent
  • Aghajari, Nushin, Institut de Biologie et Chimie des Protéines, Université de Lyon, France
organization
publishing date
type
Thesis
publication status
published
subject
keywords
carbohydrate binding modules protein engineering affinity affinity maturation phage display specificity crystallography cellulose derivatives
defense location
Lecture hall Lundmarksalen, Astronomihuset, Sölvegatan 27, Lund
defense date
2012-11-23 09:15
ISBN
978-91-7473-380-8
language
English
LU publication?
yes
id
7f17d0e4-55dd-4008-a3af-9c6a0821b9b2 (old id 3159007)
date added to LUP
2012-10-31 10:07:26
date last changed
2016-09-19 08:45:17
@misc{7f17d0e4-55dd-4008-a3af-9c6a0821b9b2,
  abstract     = {Protein-carbohydrate interactions are of central role in all living organism. In the studies presented in this thesis several strategies for engineering such interactions have been investigated. In the first study the binding affinity of a carbohydrate binding module (CBM) was improved a 100-fold through affinity maturation. Also, in that study it was discovered that all the selected, matured mutants carried a mutation in common (glutamic acid), which was shown to be responsible for the increased affinity.<br/><br>
Furthermore, the binding interaction of two different CBM was analyzed with x-ray crystallography. One of the CBM can only bind xylans, while the other binds also beta-glucans and xyloglucan. The crystal structures revealed that the binding cleft of the multi-specific CBM is flexible, permitting reorientation of side-chains to avoid steric clashes. Also the multi-specific CBM harbored an important phenylalanine that due to its chemical composition has a pi-electron surface and can interact with hydrogen atoms on the different ligands it recognizes. The specific CBM on the other hand harbor a leucine in this position and can only interact with xylans.<br/><br>
In the last study, the capacity of CBM for use as analytical tools in the characterization of the distribution pattern of cellulose derivatives was investigated. The study showed that some CBM are able to distinguish between cellulose derivatives with different substitution distribution. This is a potentially new application for CBM.<br/><br>
In all, the thesis demonstrates strategies in which carbohydrate binding proteins can be generated, improved and utilized.},
  author       = {von Schantz, Laura},
  isbn         = {978-91-7473-380-8},
  keyword      = {carbohydrate binding modules protein engineering affinity affinity maturation phage display specificity crystallography cellulose derivatives},
  language     = {eng},
  title        = {Engineering protein-carbohydrate interactions - lessons from natural and evolved carbohydrate binding modules},
  year         = {2012},
}