Role of a conserved arginine residue during catalysis in serine palmitoyltransferase

Jonathan Lowther, Guillaume Charmier, Marine C Raman, Hiroko Ikushiro, Hideyuki Hayashi, Dominic J Campopiano

Research output: Contribution to journalArticlepeer-review

Abstract

All sphingolipid-producing organisms require the pyridoxal 5'-phosphate (PLP)-dependent serine palmitoyltransferase (SPT) to catalyse the first reaction on the de novo sphingolipid biosynthetic pathway. SPT is a member of the alpha oxoamine synthase (AOS) family that catalyses a Claisen-like condensation of palmitoyl-CoA and L-serine to form 3-ketodihydrosphingosine (KDS). Protein sequence alignment across various species reveals an arginine residue, not involved in PLP binding, to be strictly conserved in all prokaryotic SPTs, the lcb2 subunits of eukaryotic SPTs and all members of the AOS family. Here we use UV-vis spectroscopy and site-directed mutagenesis, in combination with a substrate analogue, to show that the equivalent residue (R370) in the SPT from Sphingomonas wittichii is required to form the key PLP:L-serine quinonoid intermediate that condenses with palmitoyl-CoA and thus plays an essential role enzyme catalysis.
Original languageEnglish
Pages (from-to)1729-34
Number of pages6
JournalFEBS Letters
Volume585
Issue number12
Early online date18 Apr 2011
DOIs
Publication statusPublished - 2011

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