Phospholipase Cleavage of D‐ and L‐chiro‐Glycosylphosphoinositides Asymmetrically Incorporated into Liposomal Membranes

Abstract

The nature of chiro-inositol-containing inositolphosphoglycans (IPGs), reported to be putative insulin mediators, was studied by examination of the substrate specificities of the phosphatidylinositol-specific phospholipase C (PI-PLC) and the glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) by using a series of synthetic D- and L-chiro-glycosylphosphoinositides. 3-O-alpha-D-Glucosaminyl- (3) and -galactosaminyl-2-phosphatidyl-L-chiro-inositol (4), which show the maximum stereochemical similarity to the 6-O-alpha-D-glucosaminylphosphatidylinositol pseudodisaccharide motifs of GPI anchors, were synthesized and asymmetrically incorporated into phospholipid bilayers in the form of large unilamellar vesicles (LUVs). Similarly, 2-O-alpha-D-glucosaminyl- (5) and -galactosaminyl-1-phosphatidyl-D-chiro-inositol (6), which differ from the corresponding pseudodisaccharide motif of the GPI anchors only in the axial orientation of the phosphatidyl moiety, were also synthesized and asymmetrically inserted into LUVs. The cleavage of these synthetic molecules in the liposomal constructs by PI-PLC from Bacillus cereus and by GPI-PLD from bovine serum was studied with the use of 6-O-alpha-D-glucosaminylphosphatidylinositol (7) and the conserved GPI anchor structure (8) as positive controls. Although PI-PLC cleaved 3 and 4 with about the same efficiency as 7 and 8, this enzyme did not accept 5 or 6. GPI-PLD accepted both the L-chiro- (3 and 4) and the D-chiro- (5 and 6) glycosylinositolphosphoinositides. Therefore, IPGs containing L-chiro-inositol only are expected to be released from chiro-inositol-containing GPIs if the cleavage is effected by a PI-PLC, whereas GPI-PLD cleavage could result in both L-chiro- and D-chiro-inositol-containing IPGs.

The Biomedical Uses of Inositols: A Nutraceutical Approach to Metabolic Dysfunction in Aging and Neurodegenerative Diseases

“…Chelated INS-2 is an allosteric modulator of mitochondrial PDH phosphatase (PDHP) activity and promotes PDH-mediated glycogen synthesis [ 71 ]. It should be remarked that the structures of DCI-GPIs are still unknown and may not share structural similarity with MI-GPIs and differ in terms of the axial orientation of the phosphatidyl moiety, as reported by cleavage studies with synthetic DCI-GPIs ( Figure 1 C) [ 76 ].…”

ChemInform Abstract: The “Other” Inositols and Their Phosphates: Synthesis, Biology, and Medicine (with Recent Advances in myo‐Inositol Chemistry)

“…[145] Recent reviews of GPIs state that they contain myo-inositol and make no mention of d-chiro-inositol. [146] Chemically synthesized GPIs containing d-chiro-inositol are cleaved by GPI-specific phospholipase Db ut not by phosphatidylinositol-specific phospholipase C. [147] d-chiro-Inositol 1,3,4,6-tetrakisphosphate (107)i safull agonist of the inositol trisphosphate receptor in two cell lines, but the enantiomer is inactive. [83] With an IC 50 value of 1.5 mm, d-chiro-inositol 2,3,4,5-tetrakisphosphate (108)i sapotent inhibitor of Ins(3,4,5,6)P 4 1-kinase/Ins(1,3,4)P 3 5/6 kinase but its enantiomer is more than 20-fold less active.…”

The “Other” Inositols and Their Phosphates: Synthesis, Biology, and Medicine (with Recent Advances inmyo‐Inositol Chemistry)

“…[145] Recent reviews of GPIs state that they contain myo-inositol and make no mention of D-chiro-inositol. [146] Chemically synthesized GPIs containing D-chiro-inositol are cleaved by GPI-specific phospholipase D but not by phosphatidylinositol-specific phospholipase C. [147] D-chiro-Inositol 1,3,4,6-tetrakisphosphate (107) is a full agonist of the inositol trisphosphate receptor in two cell lines, but the enantiomer is inactive. [83] With an IC 50 of 1.5μM D-chiroinositol 2,3,4,5-tetrakisphosphate (108) is a potent inhibitor of Ins(3,4,5,6)P 4 1-kinase/ Ins(1,3,4)P 3 5/6 kinase but its enantiomer is more than 20-fold less active.…”

“…Recent reviews of GPIs state that they contain myo ‐inositol and make no mention of d ‐ chiro ‐inositol . Chemically synthesized GPIs containing d ‐ chiro ‐inositol are cleaved by GPI‐specific phospholipase D but not by phosphatidylinositol‐specific phospholipase C …”