N-acetylglucosamine kinase and N-acetylglucosamine 6-phosphate deacetylase in normal human erythrocytes and Plasmodium falciparum.
The major pathways of glucose metabolism in the malaria parasite, Plasmodium falciparum, have now been elucidated, and the structures and properties of parasite-specific enzymes are presently being investigated. Little is known, however, about the enzymes catalysing monosaccharide interconversions in the parasite. In the present investigation we have examined the pathway of N-acetylglucosamine catabolism which, in higher organisms, involves the following reaction sequence: N-acetylglucosamine -->N-acetylglucosamine 6-phosphate-->glucosamine 6-phosphate-->fructose 6-phosphate. Assay of the specific kinase (E.C. 2.7.1.59) catalysing the phosphorylation of the sugar showed that the enzyme is present in Plasmodium extracts as well as in normal human erythrocytes; specific activities of 7.2 and 5.3 nmol/h/mg protein were measured for the parasite and erythrocyte extracts, respectively, N-Acetylglucosamine 6-phosphate deacetylase (E.C. 3.5.1.25), catalysing the second reaction, was also detected in both normal and Plasmodium-infected erythrocytes. At 75% parasitaemia, the deacetylase activity was close to 3 times higher than that of normal control cells. The erythrocyte deacetylase was purified approximately 16,000-fold by chromatography on DE52 cellulose, chromatofocusing, and size exclusion chromatography. Attempts to purify the parasite enzyme by the same procedures were unsuccessful due to loss of activity. A partially purified erythrocyte deacetylase preparation (eluted from DE52 cellulose) had a pH optimum of 7.5, a pI of 6.0, as indicated by chromatofocusing, and a K(m) of 29 microM. In conjunction with previous investigations, the present study indicated that all three enzymes required for N-acetylglucosamine utilization are present in Plasmodium parasites as well as in normal erythrocytes.
Weidanz
J A
JA
Department of Microbiology, University of Alabama at Birmingham 35294-2170, USA.
Campbell
P
P
Moore
D
D
DeLucas
L J
LJ
Rodén
L
L
Thompson
J N
JN
Vezza
A C
AC
D000581:Amidohydrolases / Q000378:metabolism*; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D004912:Erythrocytes / Q000201:enzymology* / Q000469:parasitology; D006801:Humans; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D010963:Plasmodium falciparum / Q000201:enzymology*; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism*
D017853:Phosphotransferases (Alcohol Group Acceptor); C018716:N-acetylglucosamine kinase; D001619:beta-N-Acetylhexosaminidases; D000581:Amidohydrolases; C018717:N-acetylglucosamine-6-phosphate deacetylase