Evidence for covalent attachment of diphytanylglyceryl phosphate to the cell-surface glycoprotein of Halobacterium halobium.
In a previous study, we demonstrated the occurrence of novel proteins modified with a diphytanylglyceryl group in thioether linkage in Halobacterium halobium (Sagami, H., Kikuchi, A., and Ogura, K. (1995) J. Biol. Chem. 270, 14851-14854). In this study, we further investigated protein isoprenoid modification in this halobacterium using several radioactive tracers such as [3H]geranylgeranyl diphosphate. One of the radioactive bands observed on SDS-polyacrylamide gel electrophoresis corresponded to a periodic acid-Schiff stain-positive protein (200 kDa). Radioactive and periodic acid-Schiff stain-positive peptides (28 kDa) were obtained by trypsin digestion of the labeled proteins. The radioactive materials released by acid treatment of the peptides showed a similar mobility to dolichyl (C55) phosphate on a normal-phase thin-layer plate. However, radioactive hydrolysates obtained by acid phosphatase treatment co-migrated not with dolichol (C55-65), but with diphytanylglycerol on both reverse- and normal-phase thin-layer plates. The mass spectrum of the hydrolysate was also coincident with that of diphytanylglycerol. The partial amino acid sequences of the 28-kDa peptides were found in a fragment (amino acids 731-816) obtainable by trypsin cleavage of the known cell-surface glycoprotein of this halobacterium. These results indicate that the cell-surface glycoprotein (200 kDa) is modified with diphytanylglyceryl phosphate.
Kikuchi
A
A
Institute for Chemical Reaction Science, Tohoku University, 2-1-1, Katahira, Aobaku, Sendai 980-8577, Japan.
Sagami
H
H
Ogura
K
K
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry*; D002855:Chromatography, Thin Layer; D019617:Halobacterium salinarum / Q000737:chemistry*; D013058:Mass Spectrometry; D008562:Membrane Glycoproteins / Q000737:chemistry*; D008798:Mevalonic Acid / Q000378:metabolism; D008969:Molecular Sequence Data; D015394:Molecular Structure; D011106:Polyisoprenyl Phosphates / Q000737:chemistry; D014357:Trypsin
D001425:Bacterial Outer Membrane Proteins; D008562:Membrane Glycoproteins; D011106:Polyisoprenyl Phosphates; D014357:Trypsin; D008798:Mevalonic Acid