Structure-Affinity Relationships of Substrates for the Neutral Amino Acid Transport System in Rabbit Ileum

Abstract

The apparent affinities of various amino acids for the neutral amino acid transport system in rabbit ileum were determined by measuring the inhibition of L-methionine-14 C influx across the brush border membrane. The apparent affinity was very low for compounds lacking an a-amino group, compounds with the a-hydrogen substituted by a methyl group, D-compounds, compounds with tertiary branching in the side chain, compounds with either a positive or negative charge in the side chain, and in most cases, compounds with a hydrophilic moiety in the side chain. High apparent affinities were exhibited by compounds with unbranched carbon or carbon-sulfur side chains. Branched compounds such as valine and leucine exhibited affinities which correlate with binding of only the linear portion of the side chain. The calculated change in free energy of binding is 370 cal/mol/CH2 group which suggests the binding region for the side chain is partially hydrophobic. The affinities of families of analogues, derivatives of cysteine, methionine, serine, alanine, valine, and phenylalanine, correlate with their calculated octanol/water partition coefficients and are also correlated with apparent structural and electronic differences between families. The data permit a preliminary description of the functional geometry of the neutral amino acid transport site. The site contains a region for binding the a-amino group, a-carboxyl group, and side chain. The regions about the a-amino group and a-hydrogen are quite sterically limited. The side chain binding region is hydrophobic in nature and appears to be shallow, binding only the linear portion of branched or ring compounds.Many studies have been conducted in recent years on the relationships between the structure of substrates and their affinity for enzymes (1-8), and the results often permit deduction of general structural characteristics of the substrate binding site. Similar studies on amino acid transport systems can provide analogous information as indicated by the work of Christensen (9).

Neutral amino acid transport by marine fish intestine: role of the side chain

“…This way, which is typically diffusion, plays an important role in the transport of amino acid in fish notably at physiological concentrations (>lmmol'1-1) (Ferraris and Aheaern 1984). In Dieentrarehus labrax, we showed that this is specially the case for methionine, a probable consequence of the lipophilic properties of the side chains of this amino acid (Preston et al 1974;Brachet et al 1987). …”

“…If the K~ of the inhibitory amino acid is equal to its K, then inhibition is competitive and the two amino acids share the same transport system (Preston et al 1974;Stevens and Wright 1985).…”

“…In marine fish at least two distinct systems have been identified: the first is specific for alanine and short-chain neutral amino acids, the second is specific for imino acids and the N-methylated analogues (Vilella et al 1989;Maffia et al 1990). In mammals, numerous studies have looked into the effect of the length of this side chain on its reactivity with the amino acid transport system and it has been clearly established that the affinity of the amino acid transport mechanism is determined by the lipophilic properties of this chain, reactivity with the transport system increasing with the length of the apolar section (Preston et al 1974;Stevens and Wright 1985). The structure-affinity relationships of short-chain neutral amino acids with their specific transport systems are investigated during the present study.…”

Kinetics of Uptake of L-Leucine and Glycylsarcosine into Normal and Protein Malnourished Young Rat Jejunum

“…In this method the substrate was treated as a competitive inhibitor of its own mediated transport and the results were plotted in a form described by Preston et al (13). After extrapolation to an infinitely high concentration of "inhibitor" the uptake remaining gave the non-mediated uptake.…”

Single‐site uptake of neutral amino acids into guinea‐pig intestinal rings.

“…Nevertheless, the form of eqn. (5) is such that the points are well spaced on the graph -which is not the case of the plot proposed by Preston, Schaeffer & Curran (1974) and Matthews et al (1979) where the abscissa is the reciprocal of the inhibitor concentration. Application of the technique to the present data provided results which agreed remarkably well with those presented in Table 2 of the main text.…”