Crystal Structure of S-Ribosylhomocysteinase (LuxS) in Complex with a Catalytic 2-Ketone Intermediate

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• 作者：Rakhi Rajan ; Jinge Zhu ; Xubo Hu ; Dehua Pei ; and Charles E. Bell
• 刊名：Biochemistry
• 出版年：2005
• 出版时间：March 15, 2005
• 年：2005
• 卷：44
• 期：10
• 页码：3745 - 3753
• 全文大小：595K
• 年卷期：v.44,no.10(March 15, 2005)
• ISSN：1520-4995

文摘

S-Ribosylhomocysteinase (LuxS) is an Fe²⁺-dependent metalloenzyme that catalyzes thecleavage of the thioether bond in S-ribosylhomocysteine (SRH) to produce homocysteine (Hcys) and4,5-dihydroxy-2,3-pentanedione (DPD), the precursor of type II bacterial quorum-sensing molecule. Theproposed mechanism involves an initial metal-catalyzed aldose-ketose isomerization reaction, which resultsin the migration of the ribose carbonyl group from its C1 to C2 position and the formation of a 2-ketoneintermediate. A repetition of the isomerization reaction shifts the carbonyl group to the C3 position.Subsequent -elimination reaction at the C4 and C5 positions completes the catalytic cycle. In this work,a catalytically inactive mutant (C84A) of Co²⁺-substituted Bacillus subtilis LuxS was cocrystallized withthe 2-ketone intermediate and the structure was determined to 1.8 Å resolution. The structure reveals thatthe C2 carbonyl oxygen is directly coordinated with the metal ion, providing strong support for the proposedLewis acid function of the metal ion during catalysis. Cys-84 and Glu-57 are optimally positioned to actas general acids/bases during the isomerization and elimination reactions. In addition, Ser-6, His-11, andArg-39 are involved in substrate/ intermediate binding through hydrogen bonding interactions. The aboveconclusions are further confirmed by site-directed mutagenesis and visible absorption spectroscopic studies.