Rational protein engineering in action: the first crystal structure of a phenylalanine tRNA synthetase from Staphylococcus haemolyticus.
In this article, we describe for the first time the high-resolution crystal structure of a phenylalanine tRNA synthetase from the pathogenic bacterium Staphylococcus haemolyticus. We demonstrate the subtle yet important structural differences between this enzyme and the previously described Thermus thermophilus ortholog. We also explain the structure-activity relationship of several recently reported inhibitors. The native enzyme crystals were of poor quality--they only diffracted X-rays to 3-5A resolution. Therefore, we have executed a rational surface mutagenesis strategy that has yielded crystals of this 2300-amino acid multidomain protein, diffracting to 2A or better. This methodology is discussed and contrasted with the more traditional domain truncation approach.
Evdokimov
Artem G
AG
Pfizer Global Research & Development, Michigan Laboratories, Ann Arbor, 2800 Plymouth Road, Ann Arbor, MI 48105, USA. artem@xtals.org
Mekel
Marlene
M
Hutchings
Kim
K
Narasimhan
Lakshmi
L
Holler
Tod
T
McGrath
Teresa
T
Beattie
Bryan
B
Fauman
Eric
E
Yan
Chunhong
C
Heaslet
Holly
H
Walter
Richard
R
Finzel
Barry
B
Ohren
Jeffrey
J
McConnell
Patrick
P
Braden
Timothy
T
Sun
Fang
F
Spessard
Cindy
C
Banotai
Craig
C
Al-Kassim
Loola
L
Ma
Weijun
W
Wengender
Paul
P
Kole
Denis
D
Garceau
Norman
N
Toogood
Peter
P
Liu
Jia
J
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D018360:Crystallography, X-Ray / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016296:Mutagenesis; D010652:Phenylalanine-tRNA Ligase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D041162:Staphylococcus haemolyticus / Q000201:enzymology* / Q000235:genetics
D001426:Bacterial Proteins; D010652:Phenylalanine-tRNA Ligase