Pyridoxamine 5-phosphate oxidase and

Three enzymes play an active role in the metabolism of vitamin B6 in human erythrocytes. Pyridoxal kinase uses ATP to phosphorylate pyridoxine, pyri-doxamine, and pyridoxal. Pyridoxamine oxidase oxidizes pyridoxamine-5 -phosphate and pyridoxine-5 -phosphate to pyridoxal-5 -phosphate. The phosphatase activity produces pyridoxal from pyridoxal-5 -phosphate. The assay of the three enzymes required separation of the semicarbazone derivatives of pyridoxal-5 -phosphate and pyridoxal. The mobile phase used by Ubbink and Schnell (1988) contained 2.5% acetonitrile. Detection was by fluorescence. 

Gregory JF 3rd (1980a) Effects of epsilon-pyridoxyllysine and related compounds onliver and brain pyridoxal kinase and liver pyridoxamine (pyridoxine) 5 -phosphate oxidase. Journal of Biological Chemistry 255, 2355-9. 

The phosphorylated and non-phosphorylated forms of vitamin Bg have various physical and chemical properties. Vitamin Bs in the form of pyridoxal-5 -phosphate (PLP) and to a lesser extent, pyridoxamine-5 -phosphate (PMP), functions as a coenzyme in over 100 enzymatic reactions. All the forms of vitamin Be possess vitamin activity because they can be converted in vivo to pyridoxal. PN, PM and PL are converted to 5 -phosphate by a single kinase enzyme which in the brain and liver is most active with zinc. PNP and PMP are then converted to PLP by flavin dependent oxidase this is the reason why vitamin B2 deficiency causes a fall in available PLP (Holman 1995). Human cells can synthesize PLP from three vitamers via the Bg salvage pathway but cannot synthesize PLP de novo and must obtain it from dietary sources. 

Figure 9.1. Interconversion of the vitamin Be vitamers. Pyridoxal kinase, EC 2.7.1.38 pyridoxine oxidase, EC 1.1.1.65 pyridoxamine phosphate oxidase, EC 1.4.3.5 and pyridoxal oxidase, EC 1.1.3.12. Relative molecular masses (Mr) pyridoxine, 168.3 (hydrochloride, 205.6) pyridoxal, 167.2 pyridoxamine, 168.3 (dihydrochloride, 241.1) pyridoxal phosphate, 247.1 pyridoxamine phosphate, 248.2 and 4-pyridoxlc acid, 183.2.

Studies based on the use of an antivitamin, deoxypyri-doxine, have established that the daily requirement of the vitamin ranges between 1 and 2 mg in the human adult. A normal diet has been reported to provide 1-1.5 mg daily of the vitamin. Food appears to be the only source of the vitamin because most of the vitamin produced by the bacterial flora of the intestine is excreted in the feces, possibly after oxidation to 4-pyridoxic acid. The ingested vitamin is rapidly and completely absorbed, but the exact site of the absorption is not known. Although both pyridoxine and pyridoxamine can be excreted as such and are therefore normal constituents of human urine, part of the vitamin is oxidized to the 4-pyridoxic acid before excretion in the urine. Mammalian tissues contain at least two enzymes capable of oxidizing pyridoxine. Both enzymes seem to be flavoproteins. One attacks pyridoxine, the other attacks pyridoxine phosphate. The pyridoxine phosphate oxidase of liver has been purified 65 times. Although the enzyme was shown to act on pyridoxamine phosphate, pyridoxamine phosphate was oxidized only when the pH of the incubation mixture was raised to 10. Pyridoxine phosphate oxidase has no effect on pyridoxamine phosphate at physiological pH. 

There are several metabolic interrelationships between riboflavin and vitamin Bg. The conversion of pyridoxine or pyridoxamine phosphates to pyri-doxal phosphate is catalyzed by a flavoenzyme (pyri-doxaminephosphate oxidase EC 1.4.3.5), so that a deficiency of riboflavin may, at certain key sites, result in a secondary deficiency in Bg-dependent pathways. More evidence is needed to clarify the extent and importance of these interactions. 

---