The thrH gene product of Pseudomonas aeruginosa is a dual activity enzyme with a novel phosphoserine:homoserine phosphotransferase activity.
The thrH gene product of Pseudomonas aeruginosa has been shown to complement both homoserine kinase (thrB gene product) and phosphoserine phosphatase (serB gene product) activities in vivo. Sequence comparison has revealed that ThrH is related to phosphoserine phosphatases (PSP, EC 3.1.3.3) and belongs to the l-2-haloacid dehalogenase-like protein superfamily. We have solved the crystal structures of ThrH in the apoform and in complex with a bound product phosphate. The structure confirms an overall fold similar to that of PSP. Most of the catalytic residues of PSP are also conserved in ThrH, suggesting that similar catalytic mechanisms are used by both enzymes. Spectrophotometry-based in vitro assays show that ThrH is indeed a phosphoserine phosphatase with a K(m) of 0.207 mm and k(cat) of 13.4 min(-1), comparable with those of other PSPs. More interestingly, using high pressure liquid chromatography-based assays, we have demonstrated that ThrH is able to further transfer the phosphoryl group to homoserine using phosphoserine as the phosphoryl group donor, indicating that ThrH has a novel phosphoserine:homoserine phosphotransferase activity.
Singh
S Kumar
SK
Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
Yang
Kun
K
Karthikeyan
Subramanian
S
Huynh
Tu
T
Zhang
Xuejun
X
Phillips
Margaret A
MA
Zhang
Hong
H
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000502:physiology*; D001665:Binding Sites; D020134:Catalytic Domain; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D007700:Kinetics; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010770:Phosphotransferases / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; D010770:Phosphotransferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C120987:ThrH protein, Pseudomonas aeruginosa; C021370:homoserine kinase; D010744:Phosphoric Monoester Hydrolases; C022394:phosphoserine phosphatase