Pyridoxal kinase, pyridoxamine

This enzyme [EC 2.7.1.35] (also known as pyridoxine kinase, pyridoxamine kinase, and vitamin kinase) catalyzes the reaction of ATP with pyridoxal to produce ADP and pyridoxal 5 -phosphate. Pyridoxine, pyridoxamine, and various other derivatives can also act as substrates. 

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. 

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.

Tissue uptake of vitamin Be is again by carrier-mediated diffusion of pyridoxal (and other unphosphorylated vitamers), followed by metabolic trapping by phosphorylation. Circulating pyridoxal and pyridoxamine phosphates are hydrolyzed by extracellular alkaline phosphatase. All tissues have pyridoxine kinase activity, but pyridoxine phosphate oxidase is found mainly in the liver, kidney, and brain. 

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. 

Pyridoxal, pyridoxamine and pyfidoxine are collectively known as vitamin-B6. All three compounds are efficiently converted to the biologically active form of vitamin-B6, pyridoxal phosphate. The ATP requiring enzyme, pyridoxal kinase, catalyzes this conversion. 

The major sales form of vitamin B6 is the hydrochlorid salt of the primary alcohol pyridoxine. Another vitamin B6 form introduced in the market is the dihydrochlo-rid salt of pyridoxamine. Both vitamin B6 forms are commercially produced via various straightforward chemical synthesis routes. The biologically active cofactor is the aldehyde pyridoxal-5 -phosphate, which is derived in human or animals from the vitamin B6 forms by oxidation or transamination before or after 5 phosphorylation by pyridoxal kinases. 

Kinases catalyzing the phosphorylation of pyridox-ine, pyridoxamine, and pyridoxal have been found in microorganisms and in mammalian tissues. The kinases require zinc and magnesium, and ATP acts as phosphate donor. The affinity for the substrate varies depending on the source pyridoxine and pyridoxamine are the preferred substrates with yeast enzyme, but the mammalian enzyme has a greater affinity for pyridoxal. 

All three forms of vitamin B6 [pyridoxal, pyridoxine, and pyridoxamine] are phosphorylated by a single kinase that uses ATP as the phosphate donor. This assay describes the use of pyridoxamine as the substrate. 

The phosphorylated vitamers are dephosphorylated by membrane-bound alkaline phosphatase in the intestinal mucosa pyridoxal, pyridoxamine, and pyridoxine are all absorbed rapidly by carrier-mediated diffusion. Intestinal mucosal cells have pyridoxine kinase and pyridoxine phosphate oxidase (see Figure 9.1), so that there is net accumulation of pyridoxal phosphate by metabolic trapping. Much of the ingested pyridoxine is released into the portal circulation as pyridoxal, after dephosphorylation at the serosal surface. 

Modeling vitamin B6 metabolism is further complicated by the fact that the activity of the kinase, oxidase, and phosphatase enzymes varies between organs and species. A very simplified diagram of vitamin B6 metabolism is shown in Fig. 2. In the intestine any phosphoiylated forms are hydrolyzed. The free vitamers are readily taken up by diffusion into the intestinal wall where significant phosphorylation (Middleton, 1979) and other metabolism (Middleton, 1985) occurs. In mice small doses (up to 14 nmol) of pyridoxine (Sakurai et aL, 1988) and pyridoxamine (Sakurai et oL, 1992) were converted almost completely to pyridoxal before being released into the portal circulation. While it is dear that the intestinal microflora produce vitamin B6,... 

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. 

The term vitamin Bg refers to a group of naturally occurring pyridine derivatives represented by pyridoxine (pyridoxol, PN), pyridoxal (PL), and pyridoxamine (PM), and their phosphorylated derivatives. They are collectively referred to as vitamin Bg vitamers. The natural free forms of the vitamers could be converted to the key coenzymatic form, pyridoxal phosphate (PLP), by the action of two enzymes, a kinase and an oxidase. There are more than 140 PLP-dependent enzymatic reactions, and they are distributed in all organisms. These enzymes comprise diverse groups such as the oxidoreductases, transferases, hydrolases, lyases, and isomerases. About... 

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