SEPARATION OF NONHISTONE HIGH MOBILITY GROUP (HMG) FROM HUMAN LYMPHOCYTES BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY

The high mobility group (HMG) of nonhistone proteins have been investigated using two high performance liquid chromatographic techniques (HPLC). Reversed-phase HPLC under conditions of 50 mM triethylamine adjusted to pH 2.2 with phosphoric acid (solvent A) and 95% acetonitrile in water (solvent B) was used to separate proteins primarily on the basis of differences in the overall hydrophobicity. Size exclusion HPLC under conditions of two different solvents (A, 0.1 % trifluoroacetic acid 1FA B, 1.0% sodium dodecyl sulphate, SDS) was used to separate proteins. HMG proteins from human lymphocytes were separated into the HMG 1, HMG 2, HMG 14 and HMG 17 components. RP-HPLC is a proper method to resolve all the human lymphocyte HMG-proteins. Size exclusion HPLC was employed to resolve the HMG-protein subunits and determine their molecular weights. Ideal SE-HPLC is not capable of resolving HMG 1 from HMG 2 or HMG 14 from HM G 17 due to their molecular weight similarities. The purity of protein fractions were examined by acetic acid-urea-triton X-100 gel electrophoresis.