Acetylation of N-terminal valine of glycine N-methyltransferase affects enzyme inhibition by folate.
Native liver glycine N-methyltransferase (GNMT) is N-acetylated while the recombinant enzyme is not. We show here that acetylation of the N-terminal valine affects several kinetic parameters of the enzyme. Glycine N-methyltransferase is a regulatory enzyme mediating the availability of methyl groups by virtue of being inhibited by folate. N-acetylation does not affect the overall structure of the protein and does not affect basal enzyme activity of GNMT. Binding of both the mono- and pentaglutamate forms of 5-methyltetrahydrofolate is the same for the acetylated and non-acetylated forms of the enzyme, however the pentaglutamate form is bound more tightly than the monoglutamate form in both cases. Although binding of the folates is similar for the acetylated and non-acetylated forms of the enzyme, inhibition of enzyme activity differs significantly. The native, N-acetylated form of the enzyme shows 50% inhibition at 1.3 microM concentration of the pentaglutamate while the recombinant non-acetylated form shows 50% inhibition at 590 microM. In addition, the binding of folate results in cooperativity of the substrate S-adenosylmethionine (AdoMet), with a Hill coefficient of 1.5 for 5-methyltetrahydrofolate pentaglutamate.
Luka
Zigmund
Z
Vanderbilt University School of Medicine, Department of Biochemistry, Nashville, TN 37232, USA.
Loukachevitch
Lioudmila V
LV
Wagner
Conrad
C
D000107:Acetylation; D000494:Allosteric Regulation; D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005492:Folic Acid / Q000494:pharmacology*; D050938:Glycine N-Methyltransferase / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D008958:Models, Molecular; D011487:Protein Conformation; D051381:Rats; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014633:Valine / Q000737:chemistry
D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D005492:Folic Acid; D050938:Glycine N-Methyltransferase; C494512:Gnmt protein, rat; D014633:Valine