Synthesis of monohydroxylated inositolphosphorylceramide (IPC-C) inSaccharomyces cerevisiae requires Scs7p, a protein with both a cytochrome b5-like domain and a hydroxylase/desaturase domain

Abstract

Saccharomyces cerevisiae mutants lacking Scs7p fail to accumulate the inositolphosphorylceramide (IPC) species, IPC‐C, which is the predominant form found in wild‐type cells. Instead scs7 mutants accumulate an IPC‐B species believed to be unhydroxylated on the amide‐linked C26‐fatty acid. Elimination of the SCS7 gene suppresses the Ca2+‐sensitive phenotype of csg1 and csg2 mutants. The CSG1 and CSG2 genes are required for mannosylation of IPC‐C and accumulation of IPC‐C by the csg mutants renders them Ca2+‐sensitive. The SCS7 gene encodes a protein that contains both a cytochrome b5‐like domain and a domain that resembles the family of cytochrome b5‐dependent enzymes that use iron and oxygen to catalyse desaturation or hydroxylation of fatty acids and sterols. Scs7p is therefore likely to be the enzyme that hydroxylates the C26‐fatty acid of IPC‐C. © 1998 John Wiley & Sons, Ltd.

The Crystal Structure of an Integral Membrane Fatty Acid α-Hydroxylase

“…Based on its depth and spatial location with respect to the resolved detergent molecules in the structure, the dimetal-binding site is proposed to lie at the surface of the cytosolic side of the ER membrane. The zinc-coordinating histidine residues are located in four motifs, defined as Ia, Ib, IIa, and IIb (11). Motifs Ia and IIa are located at the C-terminal ends of TM2 and TM4, respectively, whereas motifs Ib and IIb are located in H7 and H8 within the catalytic cap domain (Fig.…”

“…The stabilized ⌬95scScs7p structure was then utilized to generate the enzyme-substrate complex. Phytoceramide, containing a 26-carbon VLCFA moiety, was used as the model substrate based on the findings of Beeler and co-workers (11). The substrate was modeled in two different orientations, with the VLCFA moiety placed in the upper channel and the LCB moiety in the lower channel and vice versa.…”

“…Although enzymes within the membrane-bound arm of the superfamily do not share extensive sequence similarity, particular features of their sequences are highly conserved. For example, distinct sequence motifs, termed "histidine boxes," contain histidine residues that are thought to coordinate two iron atoms at the active center of the enzyme (11,12). Site-directed mutagenesis of these histidines has shown that they are essential for catalysis (12).…”

“…The Saccharomyces cerevisiae endoplasmic reticulum (ER) contains five members of the membrane-bound hydroxylase/desaturase family, including sphingolipid ␣-hydroxylase (Scs7p), the yeast homolog of FA2H (11,15). In S. cerevisiae, Scs7p is responsible for adding a hydroxyl group to the ␣-carbon of the VLCFA moiety of the ceramide substrate to generate inositol phosphoceramide, one of three major sphingolipids in yeast (15).…”

Csg1p and Newly Identified Csh1p Function in Mannosylinositol Phosphorylceramide Synthesis by Interacting with Csg2p

“…Consistent with this model, Ca 2ϩ changes the sphingolipid composition by stimulating the conversion of IPC to MIPC. 2 Because Csg2p contains an EF-Ca 2ϩ -binding domain (8,10), we propose that Csg1p or Csh1p activity may be regulated by Ca 2ϩ through Csg2p.…”