Control of catalytic cycle by a pair of analogous tRNA modification enzymes.
Enzymes that use distinct active site structures to perform identical reactions are known as analogous enzymes. The isolation of analogous enzymes suggests the existence of multiple enzyme structural pathways that can catalyze the same chemical reaction. A fundamental question concerning analogous enzymes is whether their distinct active-site structures would confer the same or different kinetic constraints to the chemical reaction, particularly with respect to the control of enzyme turnover. Here, we address this question with the analogous enzymes of bacterial TrmD and its eukaryotic and archaeal counterpart Trm5. TrmD and Trm5 catalyze methyl transfer to synthesize the m1G37 base at the 3' position adjacent to the tRNA anticodon, using S-adenosyl methionine (AdoMet) as the methyl donor. TrmD features a trefoil-knot active-site structure whereas Trm5 features the Rossmann fold. Pre-steady-state analysis revealed that product synthesis by TrmD proceeds linearly with time, whereas that by Trm5 exhibits a rapid burst followed by a slower and linear increase with time. The burst kinetics of Trm5 suggests that product release is the rate-limiting step of the catalytic cycle, consistent with the observation of higher enzyme affinity to the products of tRNA and AdoMet. In contrast, the lack of burst kinetics of TrmD suggests that its turnover is controlled by a step required for product synthesis. Although TrmD exists as a homodimer, it showed half-of-the-sites reactivity for tRNA binding and product synthesis. The kinetic differences between TrmD and Trm5 are parallel with those between the two classes of aminoacyl-tRNA synthetases, which use distinct active site structures to catalyze tRNA aminoacylation. This parallel suggests that the findings have a fundamental importance for enzymes that catalyze both methyl and aminoacyl transfer to tRNA in the decoding process.
Christian
Thomas
T
Thomas Jefferson University, Department of Biochemistry and Molecular Biology, 233 South 10th Street, BLSB 220, Philadelphia, PA 19107, USA.
Lahoud
Georges
G
Liu
Cuiping
C
Hou
Ya-Ming
YM
D000604:Amino Acyl-tRNA Synthetases / Q000235:genetics / Q000378:metabolism; D000926:Anticodon; D002384:Catalysis; D020134:Catalytic Domain; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017017:Methanococcus / Q000201:enzymology; D008958:Models, Molecular; D011487:Protein Conformation; D055503:Protein Multimerization; D012343:RNA, Transfer / Q000235:genetics / Q000378:metabolism*; D012436:S-Adenosylmethionine / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D012359:tRNA Methyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D000926:Anticodon; D029968:Escherichia coli Proteins; D012436:S-Adenosylmethionine; D012343:RNA, Transfer; C489297:TrmD protein, E coli; D012359:tRNA Methyltransferases; D000604:Amino Acyl-tRNA Synthetases
Trends Biochem Sci. 2003 Jun;28(6):329-35
12826405
J Biol Chem. 2009 Jul 31;284(31):20753-62
19487703
J Biol Chem. 2001 Jun 15;276(24):20924-34
11283006
Mol Cell. 2007 Oct 26;28(2):228-39
17964262
J Biol Chem. 1992 Jul 5;267(19):13440-5
1618846
Proc Natl Acad Sci U S A. 2007 Feb 20;104(8):2620-5
17301225
Biochemistry. 2004 Jul 20;43(28):9243-55
15248782
Biochemistry. 1996 Jun 11;35(23):7308-15
8652507
FEBS Lett. 2005 Nov 7;579(27):6017-22
16226256
EMBO J. 2006 May 17;25(10):2142-54
16642040
J Mol Biol. 1997 Aug 15;271(2):209-21
9268653
Science. 1989 May 26;244(4907):986-9
2471265
Nucleic Acids Res. 2006 Feb 10;34(3):e21
16473844
J Biol Chem. 1999 May 21;274(21):14743-9
10329670
J Mol Biol. 2003 Nov 7;333(5):931-49
14583191
Biochemistry. 2005 Mar 15;44(10):3785-94
15751955
Science. 1994 Mar 11;263(5152):1432-6
8128224
BMC Mol Biol. 2006 Jul 18;7:23
16848900
Mol Syst Biol. 2006;2:2006.0008
16738554
Biochemistry. 1975 Jan 14;14(1):1-4
1109585
J Mol Biol. 2004 Jun 11;339(4):707-19
15165845
Nat Struct Biol. 1994 Sep;1(9):580-2
7634096
Nucleic Acids Res. 2005 Jan 13;33(1):307-15
15653631
J Biol Chem. 2000 Feb 18;275(7):4912-9
10671528
Genome Res. 1998 Aug;8(8):779-90
9724324
J Mol Biol. 2006 Aug 11;361(2):300-11
16843487
Nat Struct Mol Biol. 2009 Oct;16(10):1109-15
19749755
EMBO J. 1998 Sep 1;17(17):5227-37
9724658
J Biol Chem. 1987 Apr 5;262(10):4778-86
3558369
EMBO J. 2001 Jan 15;20(1-2):231-9
11226173
Mol Cell. 2007 Feb 23;25(4):531-42
17317626
EMBO J. 2001 Sep 3;20(17):4863-73
11532950
J Biol Chem. 2008 Aug 8;283(32):21997-2006
18559342
Biochemistry. 1975 Jan 14;14(1):13-8
162826
Cell. 1993 Jul 30;74(2):299-307
8343957
Annu Rev Biochem. 2000;69:617-50
10966471
EMBO J. 2003 Jun 2;22(11):2593-603
12773376
Trends Microbiol. 2006 Jan;14(1):45-54
16343907
Science. 1989 Dec 1;246(4934):1135-42
2479982
Proteins. 2008 Sep;72(4):1274-89
18384044
Nat Struct Mol Biol. 2007 Apr;14(4):272-9
17351629
J Bacteriol. 2004 Apr;186(8):2346-54
15060037
J Mol Biol. 2008 Jun 6;379(3):579-88
18466919
J Biol Chem. 2007 Sep 21;282(38):27744-53
17652090
Biochemistry. 2006 Jun 20;45(24):7463-73
16768442
J Mol Microbiol Biotechnol. 2002 Jan;4(1):71-5
11763972
J Mol Biol. 1996 Sep 6;261(5):634-45
8800212
Cell. 1994 Jan 28;76(2):357-69
8293469
J Mol Biol. 2007 Oct 26;373(3):623-32
17868690
Nat Struct Mol Biol. 2008 May;15(5):507-14
18425141
Science. 2005 Mar 25;307(5717):1969-72
15790858