Mitochondrial malic enzyme (ME2) in pancreatic islets of the human, rat and mouse  and clonal insulinoma cells.
Despite interest in malic enzyme(ME)s in insulin cells, mitochondrial malic enzyme (ME2) has only been studied with estimates of mRNA or with mRNA knockdown. Because an mRNA's level does not necessarily reflect the level of its cognate enzyme, we designed a simple spectrophotometric enzyme assay to measure ME2 activity of insulin cells by utilizing the distinct kinetic properties of ME2. Mitochondrial ME2 uses either NAD or NADP as a cofactor, has a high Km for malate and is allosterically activated by fumarate and inhibited by ATP. Cytosolic ME (ME1) and the other mitochondrial ME (ME3) use only NADP as a cofactor and have lower Kms for malate. The assay easily showed for the first time that substantial ME2 activity is present in pancreatic islets of humans, rats and mice and INS-1 832/13 cells. ME2's presence was confirmed with immunoblotting. There was no evidence that ME3 is present in these tissues.
MacDonald
Michael J
MJ
Childrens Diabetes Center, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706, USA. mjmacdon@wisc.edu
Longacre
Melissa J
MJ
Kendrick
Mindy A
MA
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D045744:Cell Line, Tumor; D002999:Clone Cells / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005650:Fumarates / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007340:Insulinoma / Q000378:metabolism*; D007515:Islets of Langerhans / Q000378:metabolism*; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008813:Mice, Inbred ICR; D008815:Mice, Inbred Strains; D008928:Mitochondria / Q000378:metabolism*; D009249:NADP / Q000737:chemistry; D051381:Rats; D017207:Rats, Sprague-Dawley; D013045:Species Specificity; D013056:Spectrophotometry, Ultraviolet / Q000379:methods; D013347:Subcellular Fractions / Q000378:metabolism
D005650:Fumarates; D009249:NADP; D000255:Adenosine Triphosphate; D008291:Malate Dehydrogenase; C030574:malate dehydrogenase (decarboxylating)
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