title: Enzymatic characterization of the pancreatic islet-specific glucose-6-phosphatase-related protein (IGRP).
abstract: Glucose is the main physiological stimulus for insulin biosynthesis and secretion by pancreatic beta-cells. Glucose-6-phosphatase (G-6-Pase) catalyzes the dephosphorylation of glucose-6-phosphate to glucose, an opposite process to glucose utilization. G-6-Pase activity in pancreatic islets could therefore be an important factor in the control of glucose metabolism and, consequently, of glucose-dependent insulin secretion. While G-6-Pase activity has been shown to be present in pancreatic islets, the gene responsible for this activity has not been conclusively identified. A homolog of liver glucose-6-phosphatase (LG-6-Pase) specifically expressed in islets was described earlier; however, the authors could not demonstrate enzymatic activity for this protein. Here we present evidence that the previously identified islet-specific glucose-6-phosphatase-related protein (IGRP) is indeed the major islet glucose-6-phosphatase. IGRP overexpressed in insect cells possesses enzymatic activity comparable to the previously described G-6-Pase activity in islets. The K(m) and V(max) values determined using glucose-6-phosphate as the substrate were 0.45 mm and 32 nmol/mg/min by malachite green assay, and 0.29 mm and 77 nmol/mg/min by glucose oxidase/peroxidase coupling assay, respectively. High-throughput screening of a small molecule library led to the identification of an active compound that specifically inhibits IGRP enzymatic activity. Interestingly, this inhibitor did not affect LG-6-Pase activity, while conversely LG-6-Pase inhibitors did not affect IGRP activity. These data demonstrate that IGRP is likely the authentic islet-specific glucose-6-phosphatase catalytic subunit, and selective inhibitors to this molecule can be obtained. IGRP inhibitors may be an attractive new approach for the treatment of insulin secretion defects in type 2 diabetes.
authors.affiliation: Millennium Pharmaceuticals, Inc, Cambridge, Massachusetts 02139, USA.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Petrolonis
authors.forename: Qing
authors.initials: Q
authors.lastname: Yang
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Tummino
authors.forename: Susan M
authors.initials: SM
authors.lastname: Fish
authors.forename: Andrea E
authors.initials: AE
authors.lastname: Prack
authors.forename: Sadhana
authors.initials: S
authors.lastname: Jain
authors.forename: Thomas F
authors.initials: TF
authors.lastname: Parsons
authors.forename: Ping
authors.initials: P
authors.lastname: Li
authors.forename: Natalie A
authors.initials: NA
authors.lastname: Dales
authors.forename: Lin
authors.initials: L
authors.lastname: Ge
authors.forename: Steven P
authors.initials: SP
authors.lastname: Langston
authors.forename: Alwin G P
authors.initials: AG
authors.lastname: Schuller
authors.forename: W Frank
authors.initials: WF
authors.lastname: An
authors.forename: Louis A
authors.initials: LA
authors.lastname: Tartaglia
authors.forename: Hong
authors.initials: H
authors.lastname: Chen
authors.forename: Suk-Bong
authors.initials: SB
authors.lastname: Hong
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002021:Buffers; D019556:COS Cells; D004396:Coloring Agents; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D016166:Free Radical Scavengers / Q000494:pharmacology; D015870:Gene Expression; D005952:Glucose-6-Phosphatase / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006943:Hyperglycemia / Q000378:metabolism / Q000503:physiopathology; D007313:Insecta; D007515:Islets of Langerhans / Q000201:enzymology*; D008099:Liver / Q000201:enzymology; D008297:Male; D008670:Metals / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D008820:Mice, Obese; D011506:Proteins / Q000235:genetics* / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D012394:Rosaniline Dyes
chemical_list: D002021:Buffers; D004396:Coloring Agents; D016166:Free Radical Scavengers; D008670:Metals; D011506:Proteins; D012333:RNA, Messenger; D012394:Rosaniline Dyes; C005095:malachite green; D005952:Glucose-6-Phosphatase; C492194:G6PC2 protein, human; C117983:G6pc2 protein, mouse; D004121:Dimethyl Sulfoxide