NADP+-dependent glutamate dehydrogenase fromAcropora formosa: purification and properties

✍ Scribed by J. Catmull; D. Yellowlees; D. J. Miller

• Publisher: Springer-Verlag
• Year: 1987
• Tongue: English
• Weight: 455 KB
• Volume: 95
• Category: Article
• ISSN: 0025-3162
• DOI: 10.1007/bf00393099

✦ Synopsis

As an initial step in our study of nitrogen metabolism in the coral/algal symbiosis we have purified glutamate dehydrogenase (EC 1.4.1.4) to homogeneity from polyp tissue of the staghorn coral Acropora formosa collected from Magnetic Island (North Queensland) in 1985-1986. The purified enzyme had a specific activity of 78 U mg -1. The native enzyme had a relative molecular weight, Mr, of 360000 (_+ 20 000), and appears to be a hexamer with subunits of Mr = 56 000 (+ 3 000). Like the enzyme from other coelenterates, the coral glutamate dehydrogenase (GDH) was absolutely specific with respect to the coenzyme substrate (NADP § and was insensitive to allosteric regulation by nucleotides; unlike other coelenterate GDHs, the coral enzyme was absolutely specific for ammonium as amino group donor in the reductive amination reaction, and major differences in kinetic properties were apparent. Linear Michaelis-Menten kinetics were observed for the substrates a-ketoglutarate, NADPH and NADP § the Km values being 0.93, 0.11 and 0.03 mM, respectively. However glutamate dehydrogenase displayed biphasic kinetics With respect to L-glutamate and ammonium, indicating two apparent Km values (18 and 81 mM for L-glutamate and 9.2 and 416 mM for ammonium). The enzyme also exhibits Scatchard plots, Hill coefficients and cooperativity indices characteristic of enzymes displaying negative cooperativity.