TY - JOUR
T1 - Crystallization of a fragment of human fibronectin
T2 - Introduction of methionine by site‐directed mutagenesis to allow phasing via selenomethionine
AU - Leahy, Daniel J.
AU - Erickson, Harold P.
AU - Aukhil, Ikramuddin
AU - Joshi, Paritosh
AU - Hendrickson, Wayne A.
PY - 1994/5
Y1 - 1994/5
N2 - Crystals of a fragment of human fibronectin encompassing the 7th through the RGD‐containing 10th type III repeats (FN7–10) have been produced with protein expressed in E. coli. The crystals are monoclinic with one molecule in the asymmetric unit and diffract to beyond 2.0 Å Bragg spacings. A mutant FN7–10 was produced in which three methionines, in addition to the single native methionine already present, have been introduced by site‐directed mutagenesis. Diffraction‐quality crystals of this mutant protein have been grown in which methionine was replaced with selenomethionine. The introduction of methionine by site‐directed mutagenesis to allow phasing from selenomethionyl‐substituted crystals is shown to be feasible by this example and is proposed as a general approach to solving the crystallographic phase problem. Strategies for selecting propitious sites for methionine mutations are discussed. © 1994 Wiley‐Liss, Inc.
AB - Crystals of a fragment of human fibronectin encompassing the 7th through the RGD‐containing 10th type III repeats (FN7–10) have been produced with protein expressed in E. coli. The crystals are monoclinic with one molecule in the asymmetric unit and diffract to beyond 2.0 Å Bragg spacings. A mutant FN7–10 was produced in which three methionines, in addition to the single native methionine already present, have been introduced by site‐directed mutagenesis. Diffraction‐quality crystals of this mutant protein have been grown in which methionine was replaced with selenomethionine. The introduction of methionine by site‐directed mutagenesis to allow phasing from selenomethionyl‐substituted crystals is shown to be feasible by this example and is proposed as a general approach to solving the crystallographic phase problem. Strategies for selecting propitious sites for methionine mutations are discussed. © 1994 Wiley‐Liss, Inc.
KW - X‐ray crystallography
KW - extracellular matrix
KW - multiwavelength anomalous diffraction (MAD)
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U2 - 10.1002/prot.340190107
DO - 10.1002/prot.340190107
M3 - Article
C2 - 8066086
AN - SCOPUS:0028332307
SN - 0887-3585
VL - 19
SP - 48
EP - 54
JO - Proteins: Structure, Function, and Bioinformatics
JF - Proteins: Structure, Function, and Bioinformatics
IS - 1
ER -