SEPARATION OF A LABELLED FRAGMENT FROM PROTEIN SYNTHESIZED IN VITRO BY THE PARS INTERMEDIA OF THE MOUSE PITUITARY GLAND

The biosynthetic activity of the pars intermedia of the mouse pituitary gland was investigated by studying the fate of labelled amino acids incorporated into relatively large-molecular-weight products (Thornton & Isherwood, 1975). It was reported that label was actively incorporated in vitro into a protein (referred to as 'labelled P1') with a molecular weight of about 75 000(Isherwood & Thornton, 1977). However, more recent observations suggest that 'labelled P1 ' is a complex between a protein and a labelled peptide. The purpose of this communication is to give a brief account of some of the findings which led to this conclusion. Full details of the incubation and cellulose acetate electrophoresis procedures are given in Isherwood & Thornton (1977). Posterior pituitary glands from male, Schofield mice were incubated in media containing [3H]leucine and [3 H] phenylalanine (50µ. / 1). In one experiment, posterior pituitary glands from ten mice were incubated with labelled amino acids for 2 h and with excess, unlabelled amino acids for lh. The tissue was homogenized in 0-1 M-HC1 with 20 unlabelled glands, and 'labelled Px ' was prepared by cellulose acetate electrophoresis. The pattern of labelling on the cellulose acetate strip was assessed by liquid scintillation counting of samples of eluate from slices of the strip, and eluate containing 'labelled Pi ' was pooled for analysis by sodium dodecyl sulphate-polyacrylamide gel electro