Methylthioadenosine Deaminase in an Alternative Quorum Sensing Pathway in Pseudomonas aeruginosa

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• 作者：Rong Guan ; Meng-Chiao Ho ; Richard F. G. Fr枚hlich ; Peter C. Tyler ; Steven C. Almo ; Vern L. Schramm
• 刊名：Biochemistry
• 出版年：2012
• 出版时间：November 13, 2012
• 年：2012
• 卷：51
• 期：45
• 页码：9094-9103
• 全文大小：636K
• 年卷期：v.51,no.45(November 13, 2012)
• ISSN：1520-4995

文摘

Pseudomonas aeruginosa possesses an unusual pathway for 5鈥?methylthioadenosine (MTA) metabolism involving deamination to 5鈥?methylthioinosine (MTI) followed by N-ribosyl phosphorolysis to hypoxanthine and 5-methylthio-伪-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鈥?254], 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 脳 10⁷ M^鈥? s^鈥? for MTA is the highest catalytic efficiency known for an MTA deaminase. 5鈥?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 IC₅₀ of 3 nM on cellular PaMTADA activity. PaMTADA is the only activity in P. aeruginosa extracts to act on MTA. MTA and 5-methylthio-伪-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 Zn²⁺, the 5鈥?methylthio group in a hydrophobic pocket, and the transition state mimic of the diazepine ring in contact with a catalytic site Glu.