Hexokinase receptor complex in hepatoma mitochondria: evidence from N,N'-dicyclohexylcarbodiimide-labeling studies for the involvement of the pore-forming protein VDAC

Biochemistry. 1986 Mar 11;25(5):1015-21. doi: 10.1021/bi00353a010.


In rapidly growing, highly glycolytic hepatoma cells as much as 65% of the total cell hexokinase is bound to the outer mitochondrial membrane [Parry, D.M., & Pedersen, P.L. (1983) J. Biol. Chem. 258, 10904-10912]. In this paper, we describe the purification to apparent homogeneity of a mitochondrial pore-forming protein from the highly glycolytic AS-30D rat hepatoma cell line. The purified protein shows a single 35 000-dalton band in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, an amino acid composition slightly more hydrophobic than that of the rat liver pore protein (also known as VDAC or mitochondrial porin), and a channel-forming activity of 136 channels min-1 (microgram of protein)-1. In addition to displaying the properties characteristic of VDAC (single-channel conductance, voltage dependence, and preference for anions), we observe that the AS-30D VDAC protein is one of only three mitochondrial proteins that bind [14C]dicyclohexylcarbodiimide (DCCD) at relatively low dosages (2 nmol of DCCD/mg of mitochondrial protein). Significantly, treatment of intact mitochondria isolated from either rat liver or the AS-30D hepatoma with DCCD results in an almost complete inhibition of their ability to binding hexokinase. Fifty percent inhibition of binding occurs at less than 2 nmol of DCCD/mg of mitochondrial protein. In contrast to DCCD, water-soluble carbodiimides are without effect on hexokinase binding. These results suggest that the pore-forming protein of tumor mitochondria forms at least part of the hexokinase receptor complex. In addition, they indicate that a carboxyl residue located within a hydrophobic region of the receptor complex may play a critical role in hexokinase binding.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acids / analysis
  • Animals
  • Carbodiimides / pharmacology*
  • Cell Line
  • Dicyclohexylcarbodiimide / pharmacology*
  • Female
  • Glycolysis
  • Hexokinase / isolation & purification
  • Hexokinase / metabolism*
  • Kinetics
  • Liver Neoplasms, Experimental / enzymology*
  • Membrane Proteins / isolation & purification
  • Membrane Proteins / metabolism*
  • Mitochondria / enzymology*
  • Molecular Weight
  • Porins*
  • Rats
  • Rats, Inbred Strains
  • Receptors, Cell Surface / isolation & purification
  • Receptors, Cell Surface / metabolism*
  • Voltage-Dependent Anion Channels


  • Amino Acids
  • Carbodiimides
  • Membrane Proteins
  • Porins
  • Receptors, Cell Surface
  • Voltage-Dependent Anion Channels
  • Dicyclohexylcarbodiimide
  • Hexokinase