The interferon-induced 67-kDa guanylate-binding protein (hGBP1) is a GTPase that converts GTP to GMP.
hGBP1 is an interferon-induced 67-kDa protein of human cells that readily binds to agarose-immobilized GTP, GDP, and GMP but not to other nucleotides. We cloned hGBP1 cDNA into a histidine-tagging vector, produced recombinant hGBP1 with 6 extra histidine residues at its N terminus in Escherichia coli, and purified this protein to near homogeneity from bacterial lysates. Purified hGBP1 hydrolyzed radiolabeled GTP but failed to hydrolyze ATP, UTP, or CTP at significant rates. Unexpectedly, the principal product of the GTP hydrolysis reaction was GMP rather than GDP. Although significant amounts of GDP were produced when the reaction was performed at 15 degrees C, GDP could not serve as substrate or as inhibitor of hGBP1. hGBP1 lacked guanylate cyclase and guanylyltransferase activity. Degradation of GTP to GMP most likely occurred via two consecutive cleavages of single phosphate groups, because pyrophosphate was not a reaction product, and because hGBP1 failed to hydrolyze GTP gamma S. In vitro modification assays with radiolabeled mevalonic acid and farnesyl pyrophosphate showed that the CaaX motif at the C terminus of hGBP1 functions as an isoprenylation signal. Thus, hGBP1 is a GTPase with novel biochemical properties that may be membrane-associated in eukaryotic cells.
Schwemmle
M
M
Abteilung Virologie, University of Freiburg, Germany.
Staeheli
P
P
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D011756:Diphosphates / Q000378:metabolism; D004926:Escherichia coli; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism / Q000494:pharmacology; D006157:Guanosine Monophosphate / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006801:Humans; D007372:Interferons / Q000494:pharmacology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016133:Polymerase Chain Reaction; D017368:Protein Prenylation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity
D017931:DNA Primers; D011756:Diphosphates; D011994:Recombinant Proteins; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006157:Guanosine Monophosphate; D006160:Guanosine Triphosphate; D007372:Interferons; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins