Combining site-specific mutagenesis and seeding as a strategy to crystallize 'difficult' proteins: the case of Staphylococcus aureus thioredoxin.
The P31T mutant of Staphylococcus aureus thioredoxin crystallizes spontaneously in space group P2(1)2(1)2(1), with unit-cell parameters a = 41.7, b = 49.5, c = 55.6 A. The crystals diffract to 2.2 A resolution. Isomorphous crystals of wild-type thioredoxin as well as of other point mutants only grow when seeded with the P31T mutant. These results suggest seeding as a valuable tool complementing surface engineering for proteins that are hard to crystallize.
Roos
Goedele
G
Brussels Center for Redox Biology, Vlaams Interuniversitair Instituut voor Biotechnologie at the Vrije Universiteit Brussel, Pleinlaan 2, 1050 Brussel, Belgium.
Brosens
Elke
E
Wahni
Khadija
K
Desmyter
Aline
A
Spinelli
Silvia
S
Wyns
Lode
L
Messens
Joris
J
Loris
Remy
R
D003460:Crystallization / Q000379:methods*; D018360:Crystallography, X-Ray; D016297:Mutagenesis, Site-Directed / Q000379:methods; D013211:Staphylococcus aureus / Q000737:chemistry* / Q000235:genetics; D013879:Thioredoxins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification
D013879:Thioredoxins