Characterization of a cell-free protein synthesizing system isolated from rat brain

Conference · Fri May 01 04:00:00 EDT 1987 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:6156240

Rajagopalan, L E; Harper, A E

The authors have characterized a cell-free preparation from rat brain that can initiate translation of endogenous mRNAs in vitro and maintain protein synthesis for at least 90 minutes at an optimum temperature of 37C. The essential component of this system is a postmitochondrial supernate (PMS) obtained by centrifuging a whole brain homogenate at 10,000 x g for 10 minutes at 4C. In the presence of phosphocreatine (PC), ATP and GTP there is active incorporation of (TVS)methionine into trichloroacetic acid precipitable protein. Incorporation is sensitive to the concentrations of PC, magnesium and potassium ions and spermidine and is inhibited 50-60% in the presence of 7-methylguanosine 5'-monophosphate, a specific inhibitor of polypeptide chain initiation. The proteolysis of brain protein that occurs when the system is incubated for more than 60 min. can be minimized by adding bovine serum albumin. The addition of 3.0 mM 5'-guanylimidodiphosphate a non-hydrolyzable analog of GTP, blocks incorporation entirely. The phosphocreatine requirement for maintaining an optimum endogenous concentration of GTP is lowered from 10.0 mM to 5.0 mM in the presence of 2.0 mM NADPH. The system that initiates protein synthesis in vitro can be used to study changes in brain protein synthesis as a result of various treatments, and the mechanisms underlying such changes.

• Research Organization: Univ. of Wisconsin, Madison
• OSTI ID: 6156240
• Report Number(s): CONF-870644-
• Conference Information: Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Journal Volume: 46:6
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACIDS
ANIMALS
ATP
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOSYNTHESIS
BODY
BRAIN
CARBOXYLIC ACIDS
CENTRAL NERVOUS SYSTEM
CHEMICAL PROPERTIES
DAYS LIVING RADIOISOTOPES
DRUGS
EVEN-ODD NUCLEI
GUANYLIC ACID
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
LIPOTROPIC FACTORS
MAMMALS
METHIONINE
NERVOUS SYSTEM
NUCLEI
NUCLEOTIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANIC SULFUR COMPOUNDS
ORGANS
PHOSPHOCREATINE
PROTEINS
RADIOISOTOPES
RATS
RODENTS
SULFUR 35
SULFUR ISOTOPES
SYNTHESIS
TRACER TECHNIQUES
VERTEBRATES