The Hexokinases from Wild-Type and Morphological Mutant Strains of Neurospora crassa

Abstract

Four hexose-phosphorylating isozymes (hexokinases a, B, y and 6) were separated by DEAEcellulose column chromatography of high-speed supernatants from Neurospora crassa wild-type strains 77a and RL3-8A. The isozymes displayed broad sugar specificity and Michaelis constants for glucose in the micromolar range. The isozymes were not inhibited by glucose 6-phosphate nor activated by citrate. Molecular weights were estimated as 95 000 by gel filtration. Glucose-specific kinases were not found.Marked quantitative variations in the relative proportion of the hexokinases were observed during growth of the fungus on minimal medium plus sucrose. Hexokinase p was the predominant form (82% of the total activity) during the period of hyphal growth. At maturation time all the isozymes were present at about the same level although a small preponderance of hexokinase fl was still evident.The relative proportions of the hexokinases did not vary when sucrose was replaced by glucose, fructose or casamino acids. When sorbose was added to the minimal medium containing sucrose, a high proportion of hexokinase p was apparent concomitantly with the well known change of growth to a colonial mode. Several morphological mutant strains were examined with respect to the hexokinase profile.Strains COI-16 and crisp showed a marked preponderance of hexokinase p. Extracts from the dapple and dingy strains lacked hexokinases a and 6. Rabbit antisera to commercial preparations of hexokinase PI and PI1 from Saccharomyces cerevisiae were able to inhibit partially the enzyme activity of Neurospora hexokinases y and 6.No effect of the antisera on hexokinases a and B could be observed.Multiple hexokinases were first recognized in the yeast Succhuromyces cerevisiae in 1961 by Trayser and Colowick [l] and by several other research groups afterwards [2,3]. The existence of two hexokinases of broad sugar specificities, designated as PI and PI1 (or A and B), and a rather specific glucokinase, seems now well established. Hexokinases PI and PI1 have been thoroughly studied by a number of investigators with respect to kinetic properties [4] and structural features [5] ; the elucidation of the amino acid sequence of one of the isozymes may be possible soon. Recent detailed genetic studies by Maitra and his group [6,7] and by Fraenkel and coworkers [8] have shown that

The Function of MoGlk1 in Integration of Glucose and Ammonium Utilization in Magnaporthe oryzae

“…In Aspergillus niger , hexokinase and glucokinase contribute similarly to the rate of glucose phosphorylation in vivo [17], [18], whereas in Neurospora crassa , four different hexokinase isoenzymes have been described [19]. In addition, double mutants in both hexokinase and glucokinase genes of Aspergillus nidulans were unable to grow on either glucose or fructose.…”

Cloning, Expression, and Characterization of the hxk-1 Gene from the White Truffle Tuber borchii Vittad.: A First Step toward Understanding Sugar Metabolism

“…All of the eukaryotic hexokinases thus cloned appear to have arisen from a common ancestor, but they have apparently all adapted to meet the specific needs of the individual cell or organism (Panneman et al, 1998). For example, in Saccharomyces cerevisiae and in Schizosaccharomyces pombe two hexokinase isoenzymes, PI and PII, coded by two different genes, have been found (Colowich, 1973;Petit et al, 1996), while in Neurospora crassa four different isoenzymes have been described (Lagos and Ureta, 1980). Among the glucose-phosphorylating enzymes, those unable to phosphorylate fructose are referred to as glucokinases (Petit and Gancedo, 1999); in Yarrowia lipolytica and in Aspergillus niger only one gene coding for hexokinase and one for glucokinase are present (Petit and Gancedo, 1999;Panneman et al, 1996).…”