In:
European Journal of Biochemistry, Wiley, Vol. 85, No. 1 ( 1978-04), p. 15-25
Abstract:
In isolated rat hepatocytes, valine is transported by a high‐ K m system ( K m = 43 mM, V = 22 nmol · mg protein −1 · min −1 ), which is non‐concentrative and energy‐independent. Valine transport proceeds equally well in both directions (influx and efflux) and is subject to accelerative exchange. By these properties, the transport system can be defined as a facilitated diffusion (passive transport). There is no evidence for valine uptake by simple (physical) diffusion (i.e. no uptake at O°C), and it is suggested that the latter takes place only in damaged or dysfunctional cells. Valine transport is temperature‐dependent, with an activation energy of 60 kJ/mol in the temperature range 30–40 °C. Below 25 °C the activation energy increases to 103 kJ/mol, possibly reflecting a phase transition in the cell membrane. Protein synthesis and protein degradation both have an activation energy of about 100 kJ/mol throughout the temperature range 10–40 °C. At 37 °C, a tracer dose of [ 14 C]valine equilibrates so rapidly across the cell membrane that it is impossible to tell whether the radioactive valine incorporated into protein is taken from an intracellular or extracellular amino acid pool. In addition, interpretation of the protein labelling pattern is complicated by endogenous protein degradation and subsequent amino acid release, which results in both isotope dilution and a gradual stimulation of protein synthesis. Since valine transport is temperature‐dependent, a slow influx or efflux of valine can be produced by lowering the incubation temperature to the range 15–25 °C. Low temperatures also reduce the disturbing influence of protein degradation, while protein synthesis still proceeds at a sufficient rate to obtain analyzable incorporation kinetics. With only a tracer dose of [ 14 C]valine in the extracellular medium, the rate of protein labelling under these conditions can be seen to follow closely the changes in the intracellular rather than in the extracellular radioactivity, both during influx and efflux of [ 14 C]valine. However, when the extracellular valine concentration is maintained several‐fold higher than the intracellular concentration, protein labelling correlates better with the extracellular than with the intracellular radioactivity. These results suggest that both the intracellular and the extracellular valine pools can directly provide precursors for protein synthesis, the relative contrib ution from each pool being proportional to the relative valine concentration in that pool, as suggested by Khairallah and co‐workers [ Biochem. J. (1974) 140 , 539–548; J. Biol. Chem. (1976) 251 , 1375–1384; Biochem. J. (1977) 162 , 257–266].
Type of Medium:
Online Resource
ISSN:
0014-2956
,
1432-1033
DOI:
10.1111/ejb.1978.85.issue-1
DOI:
10.1111/j.1432-1033.1978.tb12208.x
Language:
English
Publisher:
Wiley
Publication Date:
1978
detail.hit.zdb_id:
1398347-7
detail.hit.zdb_id:
2172518-4
SSG:
12
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