Generation and characterization of transgenic potato ( Solanum tuberosum) roots with altered levels of hexokinase

Hexokinase (HK) has been shown to highly control the rate of glycolysis in animal systems. It has been hypothesized that HK may exert a similar high level of control within plant systems, especially during periods of O ₂ limitations. In order to quantify the level of control transgenic potato roots were created with altered levels of HK protein and enzyme activity. A HK (SCHK) cDNA clone of Solanum chacoense was sequenced and analyzed. The SCHK amino acid sequence is nearly identical to S. tuberosum HK2. The sequence also shows a high level of similarity to other plant HKs, when compared to non-plant eukaryotic HKs the sequences share similarities in conserved binding domains. Transforming potato with SCHK using sense and antisense constructs in an Agrobacterium rhizogenes transformation system created transgenic roots. The resulting root lines had a range of HK protein from 0.2 to 4-fold that of the control lines, as determined by immunodetection. This alteration in protein level corresponded to an increase in phosphorylation activity using both glucose and fructose as substrates. A HK activity range of 0.6 to 6.4-fold of the control lines was observed when using glucose (GK) as substrate and 0.7 to 4.5-fold range of activity when using fructose (FK). This range in protein and HK activity is broad enough for use in determining the degree of control that HK exerts on the rate of potato root glycolysis.