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Transport of L‐alanine in cultured human fibroblasts: Evidence for two kinetically distinguishable systems
Author(s) -
Feneant M.,
Moatti N.,
Lemonnier F.,
Lemonnier A.
Publication year - 1981
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041090105
Subject(s) - alanine , chemistry , substrate (aquarium) , kinetics , enzyme kinetics , enzyme , stereochemistry , biochemistry , amino acid , biology , active site , physics , ecology , quantum mechanics
Abstract The transport of L‐alanine in human diploid fibroblasts was investigated. Transport measurements were performed on subcultures between the third and eighth passages with subconfluent cells growing on glass coverslips. Kinetic analysis of approximate initial rates of transport at substrate concentrations from 0.05 to 10 mmole/liter indicate the presence of two distinguishable systems. The high affinity system has a K m of 0.24 mmole/liter and a V max of 6.4 nmole/100 μg protein/2 min. For the low affinity system, the contribution of the high affinity system to the uptake must absolutely be taken into account. The K m and V max values, obtained by using a computer program, are a K m of 15.0 mmole/liter and a V max of 14.7 nmole/100 μg protein/2 min. For alanine concentrations below 1 mmole/liter, the contribution of the Na + ‐independent uptake is less than 10%, and the kinetic constants of the high affinity system are in the same range if this contribution is taken into account. On the contrary the influence of a diffusion‐like process is more significant on the low affinity system whose K m is about 49 mmole/liter after subtraction of the Na + ‐independent uptake from the experimental velocities. Inhibition studies were performed with NCH 3 ‐alanine. They permitted us first to confirm the existence of system A in cultured human fibroblasts in agreement with two recent works and second to show how this system contributes to L‐alanine uptake. This contribution seems very small in low concentrations but it rises as the concentrations increase.