title: Identification of a mitochondrial-binding site on the N-terminal end of hexokinase II.
abstract: Hexokinase II (HKII) is responsible for the first step in the glycolysis pathway by adding a phosphate on to the glucose molecule so it can proceed down the pathway to produce the energy for continuous cancer cell growth. Tumour cells overexpress the HKII enzyme. In fact, it is the overexpression of the HKII enzyme that makes the diagnosis of cancer possible when imaged by positron emission tomography (PET). HKII binds to the voltage-dependent anion channel (VDAC) located on the mitochondrial outer membrane (MOM). When bound to the MOM, HKII is blocking a major cell death pathway. Thus, HKII is responsible for two characteristics of cancer cells, rapid tumour growth and inability of cancer cells to undergo apoptosis. One method to identify novel compounds that may interfere with the HKII-VDAC-binding site is to create a molecular model using the crystal structure of HKII. However, the amino acid(s) responsible for HKII binding to VDAC are not known. Therefore, a series of truncations and point mutations were made to the N-terminal end of HKII to identify the binding site to VDAC. Deletions of the first 10 and 20 amino acids indicated that important amino acid(s) for binding were located within the first 10 amino acids. Next, a series of point mutations were made within the first 10 amino acids. It is clear from the immunofluorescence images and immunoblot results that mutating the fifth amino acid from histidine to proline completely abolished binding to the MOM.
authors.affiliation: Department of Otolaryngology, University of Florida, Gainesville, FL 32610, USA.
authors.forename: Nadezda
authors.initials: N
authors.lastname: Bryan
authors.affiliation: Department of Otolaryngology, University of Florida, Gainesville, FL 32610, USA raisch@ufl.edu.
authors.forename: Kevin P
authors.initials: KP
authors.lastname: Raisch
mesh_terms: D001665:Binding Sites; D002460:Cell Line; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006593:Hexokinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000378:metabolism; D009154:Mutation; D017354:Point Mutation; D050994:Voltage-Dependent Anion Channels / Q000378:metabolism
chemical_list: D050994:Voltage-Dependent Anion Channels; D049452:Green Fluorescent Proteins; D006593:Hexokinase
keywords: cancer; glycolysis; hexokinase II; mintochondrial-binding site; mutations