High impact information on HXK2

• Deletion of HEX1 in N. crassa eliminates Woronin bodies from the cytoplasm and results in hyphae that exhibit a cytoplasmic-bleeding phenotype in response to cell lysis [1].
• Here we purify the Woronin body from Neurospora crassa and isolate Hex1, a new protein containing a consensus sequence known as peroxisome-targeting signal-1 (PTS1) [1].
• We show that Hex1 is localized to the matrix of the Woronin body by immunoelectron microscopy, and that a green fluorescent protein- (GFP-)Hex1 fusion protein is targeted to yeast peroxisomes in a PTS1- and peroxin-dependent manner [1].
• In contrast, expression of LexA-Reg1p containing mutations at phenylalanine in the putative PP-1C-binding site motif (K/R)(X)(I/V)XF was unable to rescue Hxk2p dephosphorylation in intact yeast or restore Hxk2p phosphatase activity [2].
• Mixed peptide sequencing identified these proteins as hexokinase II (Hxk2p) and the E1alpha subunit of pyruvate dehydrogenase [2].

Biological context of HXK2

• An HXK2 mutation encoding a serine-to-alanine change at position 15 [HXK2 (S15A)] was to clarify the in vivo function of the phosphorylation of hexokinase PII [3].
• Principal components analysis of the summed fractional labelling data show that deleting the genes HXK2 and GRR1 results in similar phenotype at the fluxome level, with a partial alleviation of glucose repression on the respiratory metabolism [4].
• In this study, suppressor mutations of the growth defect of a tps1delta hxk1delta double mutant on fructose were isolated and identified as novel HXK2 alleles [5].
• The predicted amino acid sequence of the HXK2 gene product has significant homology to the conserved catalytic domain of mammalian and yeast protein kinases [6].
• These HXK2::lacZ fusions differ in the amount of the HXK2 gene (encoding hexokinase 2 isoenzyme) that is fused to the lacZ reporter gene [7].

Anatomical context of HXK2

• The HEX1 gene product was functional in S. cerevisiae cells and was targeted to the periplasm [8].

Associations of HXK2 with chemical compounds

• A series of deletions of this HXK2 coding region were constructed and fused upstream of a minimal CYC1::lacZ promoter. beta-Galactosidase activities on glucose or ethanol growth yeast calls revealed that two different regulatory elements are present in this DNA region [7].
• All strains with HXK2 showed repression in growth on glucose and the others did not [9].
• However, the ggs1 delta, hxk2 delta double mutant still displayed a high level of glucose-6-phosphate as well as the rapid appearance of free intracellular glucose [10].
• Mig1 and Hxk2 interacted in vivo in a yeast two-hybrid assay and in vitro in immunoprecipitation and glutathione S-transferase pull-down experiments [11].
• Expression of hxk2+ was highest during growth in glycerol [12].

Physical interactions of HXK2

• Mediator factor Med8p interacts with the hexokinase 2: implication in the glucose signalling pathway of Saccharomyces cerevisiae [13].
• We conclude that Hxk2 operates by interacting with Mig1 to generate a repressor complex located in the nucleus of S. cerevisiae during growth in glucose medium [11].
• Furthermore, we demonstrate using gel mobility shift analysis that Hxk2 participates in DNA-protein complexes with cis-acting regulatory elements of the SUC2 gene promoter [14].
• The surface area of the hexokinase A-glucose complex exposed to solvent is smaller than that of native hexokinase B [15].

Enzymatic interactions of HXK2

• We reduced hexokinase activity in ggs1 strains by deleting the gene HXK2 encoding hexokinase PII [10].
• Mutants of gene HEX1 had a reduced hexose phosphorylating activity on all media whereas those of gene HEX2 had elevated levels but only in glucose grown cells [16].

Regulatory relationships of HXK2

• Using the set of six hxk2 mutants it was shown that there is a good correlation between the glucose-induced cAMP signal and in vivo hexokinase activity [5].
• Rgt1 represses HXK2 expression by binding specifically to the motif (CGGAAAA) located at -395 bp relative to the ATG translation start codon in the HXK2 promoter [17].
• Here we report that Hxk2 has a glucose-regulated nuclear localization and that Mig1, a transcriptional repressor responsible for glucose repression of many genes, is required to sequester Hxk2 into the nucleus [11].
• Tpk3 and Snf1 protein kinases regulate Rgt1 association with Saccharomyces cerevisiae HXK2 promoter [18].
• An hxk2-delta 1::URA3 mutant did not detectably express Hxt2p in ethanol or galactose, but an ssn6-delta9 mutant did highly express Hxt2p in both carbon sources [19].

Other interactions of HXK2

• The hxk2 single mutant, as well as the double mutant, failed to show catabolite repression in all three systems, while the hxk1 null mutation had little or no effect on catabolite repression [20].
• Components of the glucose repression pathway (Hxk2p and Reg1p) are also required for generation of the high-level glucose induction signal for expression of the HXT1 gene [21].
• We conclude that Hxk2p operates through the MED8 site, by interacting with Med8p, in the glucose signal transduction pathway of Saccharomyces cerevisiae [13].
• The expression of CAT8-lacZ and SIP4-lacZ has been also measured in mig1, tup1 or hxk2 mutants, partially refractory to catabolite repression [22].
• The glucose inhibition pathway requires HXK2, REG1, and GSF1 and appears to overlap upstream with the glucose repression pathway [23].

Analytical, diagnostic and therapeutic context of HXK2

• Analysis by atomic force microscopy of Med8 binding to cis-acting regulatory elements of the SUC2 and HXK2 genes of saccharomyces cerevisiae [24].
• The change in enthalpy and rate constants for the reactions of yeast hexokinase isozymes, PI (Hxk1) and PII (Hxk2), was determined at pH 7.6 and 25 degrees C by isothermal titration calorimetry [25].
• Sequencing a protein by x-ray crystallography. I. Interpretation of yeast hexokinase B at 2.5 A resolution by model building [26].
• In the present study we employed site-directed mutagenesis of amino acids important for the binding of sugar and ATP, for efficient phosphoryl transfer and for the closure of the substrate-binding cleft, to obtain an insight into the structural requirements of Hxk2 for sugar-induced signalling [27].
• The binding of D-glucose to hexokinase PII at 25 degrees C and pH 8.7 has been investigated by a combination of ultrasonic velocimetry, high precision densimetry, and fluorescence spectroscopy [28].

References