Two Arabidopsis Genes (IPMS1 and IPMS2) Encode Isopropylmalate Synthase, the Branchpoint Step in the Biosynthesis of Leucine
2007
de Kraker, Jan-Willem | Luck, Katrin | Textor, Susanne | Tokuhisa, James G. | Gershenzon, Jonathan
Heterologous expression of the Arabidopsis (Arabidopsis thaliana) IPMS1 (At1g18500) and IPMS2 (At1g74040) cDNAs in Escherichia coli yields isopropylmalate synthases (IPMSs; EC 2.3.3.13). These enzymes catalyze the first dedicated step in leucine (Leu) biosynthesis, an aldol-type condensation of acetyl-coenzyme A (CoA) and 2-oxoisovalerate yielding isopropylmalate. Most biochemical properties of IPMS1 and IPMS2 are similar: broad pH optimum around pH 8.5, Mg²⁺ as cofactor, feedback inhibition by Leu, Km for 2-oxoisovalerate of approximately 300 μM, and a Vmax of approximately 2 x 10³ μmol min⁻¹ g⁻¹. However, IPMS1 and IPMS2 differ in their Km for acetyl-CoA (45 μM and 16 μM, respectively) and apparent quaternary structure (dimer and tetramer, respectively). A knockout insertion mutant for IPMS1 showed an increase in valine content but no changes in Leu content; two insertion mutants for IPMS2 did not show any changes in soluble amino acid content. Apparently, in planta each gene can adequately compensate for the absence of the other, consistent with available microarray and reverse transcription-polymerase chain reaction data that show that both genes are expressed in all organs at all developmental stages. Both encoded proteins accept 2-oxo acid substrates in vitro ranging in length from glyoxylate to 2-oxohexanoate, and catalyze at a low rate the condensation of acetyl-CoA and 4-methylthio-2-oxobutyrate, i.e. a reaction involved in glucosinolate chain elongation normally catalyzed by methylthioalkylmalate synthases. The evolutionary relationship between IPMS and methylthioalkylmalate synthase enzymes is discussed in view of their amino acid sequence identity (60%) and overlap in substrate specificity.
اظهر المزيد [+] اقل [-]الكلمات المفتاحية الخاصة بالمكنز الزراعي (أجروفوك)
المعلومات البيبليوغرافية
تم تزويد هذا السجل من قبل National Agricultural Library