Structure and alternative splicing of the ketohexokinase gene

Abstract

Ketohexokinase (fructokinase, KHK) catalyses the phosphorylation of fructose to fructose-1-phosphate. It thereby initiates the intracellular catabolism of a large proportion of dietary carbohydrate. Although found at high level in liver, renal cortex and small intestine, fructokinase activity has also been known for many years to be present at lower levels in most other tissues. We previously found that there appeared to be two isoforms of human KHK, and have now investigated the molecular basis for this in human, rat and mouse. Cloning of the human KHK gene, on chromosome 2p23.2-2p23.3, shows that it has nine exons, spanning 14 kb. An intragenic duplication has resulted in two similar 135-bp exons (designated 3a and 3c), separated by a short intron. Exon 3a and exon 3c are mutually exclusively spliced into KHK mRNA. This exonϪintron structure and the pattern of alternative splicing are conserved in both the rat and mouse, suggesting distinct conserved functions for the two KHK isoforms. The alternative splicing is also tissue specific, since in both rat and human, tissues expressing high levels of KHK (liver, kidney and duodenum) utilise exclusively the 3c exon, while other tissues use only 3a. Furthermore, comparison of human foetal and adult tissues indicates a developmental splicing shift from use of exon 3a to exon 3c.Keywords : fructokinase; ketohexokinase; alternative splicing ; isozyme.Ketohexokinase (fructokinase, KHK) catalyses the phosphorylation of fructose to fructose-1-phosphate (F1P). The enzyme can also phosphorylate a variety of other furanose sugars, and the substrate requirements for hepatic KHK have been defined in detail [1]. Dietary fructose (including the large quantity derived from sucrose hydrolysis) is thereby entered into a specialised catabolic pathway involving aldolase B and triokinase. This pathway metabolises fructose to the glycolytic intermediate glyceraldehyde-3-phosphate, in the process bypassing the important regulatory phosphofructokinase/fructose bisphosphatase step of glycolysis. KHK is most abundant in the liver, where it constitutes between 0.04% and 0.6 % of total protein [1,2]. The enzyme has been purified from human [3], bovine [1] and rat liver [2]. In its active form, it is a dimer [3] and requires K ϩ and ATP for activity. The physiological substrate for hepatic KHK is D-fructose, derived from dietary fructose, sucrose, or sorbitol, but it can also phosphorylate D-xylulose [1,3].Although most abundant in the liver, KHK is also found in kidney, small intestine and pancreas [2] and at much lower levels in heart, brain and muscle [3]. The distribution of highexpressing tissues is generally consistent with a major role in

Properties of Normal and Mutant Recombinant Human Ketohexokinases and Implications for the Pathogenesis of Essential Fructosuria

“…Those high-expressing tissues listed above contain KHK-C mRNA, with little or no KHK-A. In other tissues, KHK-A mRNA is present at low level, but KHK-C cannot be detected (12). There is also a shift from KHK-A to KHK-C in the liver and kidney during fetal development, which correlates with earlier observations that fructokinase activity is very low in fetal liver (16).…”

“…The human KHK gene on chromosome 2p23 has nine exons spanning 14 kb (12). Two exons (referred to as 3a and 3c, each 135 bp long) arose by an intragenic duplication and are mutually exclusively spliced into mRNA.…”

“…The biochemically well-characterized KHK-C, usually referred to as fructokinase, is encoded by the mRNA that includes exon 3c. It is abundant in liver, renal cortex, and small intestine and also is present in the pancreatic islet (12,13).…”

Opposing effects of fructokinase C and A isoforms on fructose-induced metabolic syndrome in mice

“…Fructokinase exists in two alternatively spliced isoforms consisting of fructokinase C (KHK-C) and fructokinase A (KHK-A) differing in exon 3 (19,20). KHK-C is expressed primarily in the liver, kidney, and intestines, whereas KHK-A is more ubiquitous (21).…”

Bioactivity-guided identification of botanical inhibitors of ketohexokinase

“…Due to alternative splicing, the expressions of the isoforms are tissue dependent. While KHKA is expressed at low levels in most tissues, high levels of KHKC are primarily expressed in the liver, kidneys, and small intestine [24,25]. Notably, only recombinant human KHKC, but not KHKA, was capable of rapidly metabolizing fructose and causing acute depletion of hepatic ATP [11].…”