Subcellular Particles: A Symposium Held during the Meeting of the Society of General Physiologists at the Marine Biological Laboratory, Woods Hole, Massachusetts, June 9-11, 1958

By Teru Hayashi | Go to book overview

Intermediate Reactions in Protein Synthesis1,2,3

MARY L. STEPHENSON, LISELOTTE I. HECHT, JOHN W. LITTLEFIELD, ROBERT B. LOFTFIELD AND PAUL C. ZAMECNIK

Massachusetts General Hospital Boston, Massachusetts

SINCE ACCEPTING THE PLEASANT INVITATION to participate in a symposium on subcellular particles we wondered what to present in the time allotted to us. Recently several reviews have covered the field of protein synthesis and the critical role of the ribonucleoprotein particle as the initial site of incorporation of amino acid into protein has been well established (32, 8, 20, 10). For these reasons we decided that a presentation of our current studies dealing primarily with the non-particulate enzymatic reactions preceding peptide bond formation would be timely and not out of place.

The present discussion of the problem is not on the whole a historical approach, but rather is designed to point out the essential factors and possible steps involved. Most of the experiments described herein have been carried out with mammalian cell systems and deal predominantly with the mechanism of the incorporation of amino acids into uncharacterized proteins rather than with the net synthesis of one or more specific proteins.

The initial site of the incorporation of amino acid into protein has been shown by several independent investigators to be in the microsome fraction (5, 25, 28, 30, 1, 36). In a representative experiment the amino acid was administered to the whole animal, after which, at various time intervals, the livers or other organs were removed, homogenized, and separated into fractions by differential centrifugation. The proteins in the microsomal fraction were the most rapidly labeled.

As is well known, the microsomal fraction refers to the pellet obtained after centrifugation of the 15,000 g supernatant of a homogenate for one or two hours at 15,000 g. The bulk of the cytoplasmic ribonucleic acid (RNA) is concentrated in this fraction (9, 7, 2). Two fractions can be obtained after treatment of microsomes with sodium deoxycholate (37). One is an insoluble fraction consisting of dense spherical particles with a diameter of about 150-250 A, which contain

____________________
1
This investigation was supported by grants-in-aid from the U. S. Atomic Energy Commission, the Public Health Service and the American Cancer Society.
2
Publication no. 926 of the Harvard Cancer Commission.
3
From the John Collins Warren Laboratories of the Collis P. Huntington Memorial Hospital of Harvard University at the Massachusetts General Hospital.

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