Header

UZH-Logo

Maintenance Infos

Crystal structures of designed armadillo repeat proteins: Implications of construct design and crystallization conditions on overall structure


Reichen, Christian; Madhurantakam, Chaithanya; Plückthun, Andreas; Mittl, Peer R E (2014). Crystal structures of designed armadillo repeat proteins: Implications of construct design and crystallization conditions on overall structure. Protein Science, 23(11):1572-1583.

Abstract

Designed armadillo repeat proteins (dArmRP) are promising modular proteins for the engineering of binding molecules that recognize extended polypeptide chains. We determined the structure of a dArmRP containing five internal repeats and 3rd generation capping repeats in three different states by X-ray crystallography: without N-terminal His6 -tag and in the presence of calcium (YM5 A/Ca(2+) ), without N-terminal His6 -tag and in the absence of calcium (YM5 A), and with N-terminal His6 -tag and in the presence of calcium (His-YM5 A/Ca(2+) ). All structures show different quaternary structures and superhelical parameters. His-YM5 A/Ca(2+) forms a crystallographic dimer, which is bridged by the His6 -tag, YM5 A/Ca(2+) forms a domain-swapped tetramer, and only in the absence of calcium and the His6 -tag, YM5 A forms a monomer. The changes of superhelical parameters are a consequence of calcium binding, because calcium ions interact with negatively charged residues, which can also participate in the modulation of helix dipole moments between adjacent repeats. These observations are important for further optimizations of dArmRPs and provide a general illustration of how construct design and crystallization conditions can influence the exact structure of the investigated protein.

Abstract

Designed armadillo repeat proteins (dArmRP) are promising modular proteins for the engineering of binding molecules that recognize extended polypeptide chains. We determined the structure of a dArmRP containing five internal repeats and 3rd generation capping repeats in three different states by X-ray crystallography: without N-terminal His6 -tag and in the presence of calcium (YM5 A/Ca(2+) ), without N-terminal His6 -tag and in the absence of calcium (YM5 A), and with N-terminal His6 -tag and in the presence of calcium (His-YM5 A/Ca(2+) ). All structures show different quaternary structures and superhelical parameters. His-YM5 A/Ca(2+) forms a crystallographic dimer, which is bridged by the His6 -tag, YM5 A/Ca(2+) forms a domain-swapped tetramer, and only in the absence of calcium and the His6 -tag, YM5 A forms a monomer. The changes of superhelical parameters are a consequence of calcium binding, because calcium ions interact with negatively charged residues, which can also participate in the modulation of helix dipole moments between adjacent repeats. These observations are important for further optimizations of dArmRPs and provide a general illustration of how construct design and crystallization conditions can influence the exact structure of the investigated protein.

Statistics

Citations

7 citations in Web of Science®
8 citations in Scopus®
Google Scholar™

Altmetrics

Downloads

0 downloads since deposited on 10 Dec 2014
0 downloads since 12 months

Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Department of Biochemistry
07 Faculty of Science > Department of Biochemistry
Dewey Decimal Classification:570 Life sciences; biology
Language:English
Date:November 2014
Deposited On:10 Dec 2014 15:59
Last Modified:05 Apr 2016 18:36
Publisher:Wiley-Blackwell Publishing, Inc.
ISSN:0961-8368
Free access at:Publisher DOI. An embargo period may apply.
Publisher DOI:https://doi.org/10.1002/pro.2535
PubMed ID:25132085

Download