Methylthioadenosine deaminase in an alternative quorum sensing pathway in Pseudomonas aeruginosa.
Pseudomonas aeruginosa possesses an unusual pathway for 5'-methylthioadenosine (MTA) metabolism involving deamination to 5'-methylthioinosine (MTI) followed by N-ribosyl phosphorolysis to hypoxanthine and 5-methylthio-α-d-ribose 1-phosphate. The specific MTI phosphorylase of P. aeruginosa has been reported [Guan, R., Ho, M. C., Almo, S. C., and Schramm, V. L. (2011) Biochemistry 50, 1247-1254], and here we characterize MTA deaminase from P. aeruginosa (PaMTADA). Genomic analysis indicated the PA3170 locus to be a candidate for MTA deaminase (MTADA). Protein encoded by PA3170 was expressed and shown to deaminate MTA with 40-fold greater catalytic efficiency for MTA than for adenosine. The k(cat)/K(m) value of 1.6 × 10(7) M(-1) s(-1) for MTA is the highest catalytic efficiency known for an MTA deaminase. 5'-Methylthiocoformycin (MTCF) is a 4.8 pM transition state analogue for PaMTADA but causes no significant inhibition of human adenosine deaminase or MTA phosphorylase. MTCF is permeable to P. aeruginosa and exhibits an IC(50) of 3 nM on cellular PaMTADA activity. PaMTADA is the only activity in P. aeruginosa extracts to act on MTA. MTA and 5-methylthio-α-d-ribose are involved in quorum sensing pathways; thus, PaMTADA is a potential target for quorum sensing. The crystal structure of PaMTADA in complex with MTCF shows the transition state mimic 8(R)-hydroxyl group in contact with a catalytic site Zn(2+), the 5'-methylthio group in a hydrophobic pocket, and the transition state mimic of the diazepine ring in contact with a catalytic site Glu.
Department of Biochemistry, Albert Einstein College of Medicine, Yeshiva University, 1300 Morris Park Avenue, Bronx, New York 10461, United States.
Rong
R
Guan
Meng-Chiao
MC
Ho
Richard F G
RF
Fröhlich
Peter C
PC
Tyler
Steven C
SC
Almo
Vern L
VL
Schramm
D000243:Adenosine Deaminase / Q000378:metabolism; D000595:Amino Acid Sequence; D003070:Coformycin / Q000031:analogs & derivatives / Q000494:pharmacology; D018360:Crystallography, X-Ray; D003839:Deoxyadenosines / Q000378:metabolism*; D006801:Humans; D008025:Ligases; D008778:Methylthioinosine / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009700:Nucleoside Deaminases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D053038:Quorum Sensing; D016415:Sequence Alignment; D013379:Substrate Specificity; D013872:Thionucleosides / Q000378:metabolism*
D003839:Deoxyadenosines; D013872:Thionucleosides; D003070:Coformycin; D008778:Methylthioinosine; C008500:5'-methylthioadenosine; D009700:Nucleoside Deaminases; C578386:methylthioadenosine deaminase, Pseudomonas aeruginosa; C548482:ADA protein, human; D000243:Adenosine Deaminase; D008025:Ligases; C449816:N-acylhomoserine lactone synthase