Kinetic Manifestation of Processivity during Multiple Methylations Catalyzed by SET Domain Protein Methyltransferases
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- 作者：Lynnette M. A. Dirk ; E. Megan Flynn ; Kevin Dietzel ; Jean-Franç ; ois Couture ; Raymond C. Trievel ; Robert L. Houtz
- 刊名：Biochemistry
- 出版年：2007
- 出版时间：March 27, 2007
- 年：2007
- 卷：46
- 期：12
- 页码：3905 - 3915
- 全文大小：552K
- 年卷期：v.46,no.12(March 27, 2007)
- ISSN：1520-4995
文摘
Processive versus distributive methyl group transfer was assessed for pea Rubisco large subunitmethyltransferase, a SET domain protein lysine methyltransferase catalyzing the formation of trimethyllysine-14 in the large subunit of Rubisco. Catalytically competent complexes between an immobilizedform of des(methyl) Rubisco and Rubisco large subunit methyltransferase were used to demonstrate enzymerelease that was co-incident with and dependent on formation of trimethyllysine. Catalytic rate constantsdetermined for formation of trimethyllysine were considerably lower (~10-fold) than rate constantsdetermined for total radiolabel incorporation from [3H-methyl]-S-adenosylmethionine. Double-reciprocalvelocity plots under catalytic conditions favoring monomethyllysine indicated a random or ordered reactionmechanism, while conditions favoring trimethyllysine suggested a hybrid ping-pong mechanism. Theseresults were compared with double-reciprocal velocity plots and product analyses obtained for HsSET7/9(a monomethyltransferase) and SpCLR4 (a dimethyltransferase) and suggest a predictive ability of double-reciprocal velocity plots for single versus multiple methyl group transfers by SET domain protein lysinemethyltransferases. A model is proposed for SET domain protein lysine methyltransferases in which initialbinding of polypeptide substrate and S-adenosylmethionine is random, with polypeptide binding followedby deprotonation of the 
lon.gif" BORDER=0 >-amine of the target lysyl residue and subsequent methylation. Following methylgroup transfer, S-adenosylhomocysteine and monomethylated polypeptide dissociate from monomethyltransferases, but di- and trimethyltransferases begin a successive and catalytically obligatory deprotonationof enzyme-bound methylated lysyl intermediates, which along with binding and release of S-adenosylmethionine and S-adenosylhomocysteine is manifested as a hybrid ping-pong-like reactionmechanism.
lon.gif" BORDER=0 >-amine of the target lysyl residue and subsequent methylation. Following methylgroup transfer, S-adenosylhomocysteine and monomethylated polypeptide dissociate from monomethyltransferases, but di- and trimethyltransferases begin a successive and catalytically obligatory deprotonationof enzyme-bound methylated lysyl intermediates, which along with binding and release of S-adenosylmethionine and S-adenosylhomocysteine is manifested as a hybrid ping-pong-like reactionmechanism.