Uptake of phenylalanine into isolated barley vacuoles is driven by both tonoplast adenosine triphosphatase and pyrophosphatase: evidence for a hydrophobic L-amino acid carrier system

The uptake of phenylalanine was studied with vacuole isolated from barley mesophyll protoplasts. The phenylalanine transport exhibited saturation kinetics with apparent Km-values of 1.2 to 1.4 millimolar for ATP- or PPi-driven uptake; Vmax apparent was 120 to 140 nanomoles Phe per milligram of chlor...

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Bibliographic Details
Published in:Plant physiology (Bethesda) 1989-04, Vol.89 (4), p.1388-1393
Main Authors: Homeyer, U, Litek, K, Huchzermeyer, B, Schultz, G
Format: Article
Language:English
Subjects:
actividad enzimatica
activite enzymatique
Adenosine triphosphatases
adenosine triphosphate
adenosintrifosfato
alanina
alanine
Amino acids
Barley
Enzymes
enzymic activity
fenilalanina
hidrolasas
hordeum vulgare
hydrolase
hydrolases
Hydrolysis
leucina
leucine
Nucleotides
nutrient transport
phenylalanine
Plants
Protoplasts
Tonoplast
transport des substances nutritives
transporte de nutrientes
triptofano
tryptophan
tryptophane
vacuola
vacuole
Vacuoles
valina
valine
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Summary:The uptake of phenylalanine was studied with vacuole isolated from barley mesophyll protoplasts. The phenylalanine transport exhibited saturation kinetics with apparent Km-values of 1.2 to 1.4 millimolar for ATP- or PPi-driven uptake; Vmax apparent was 120 to 140 nanomoles Phe per milligram of chlorophyll per hour (1 milligram of chlorophyll corresponds to 5 X 10(6) vacuoles). Half-maximal transport rates driven with ATP or PPi were reached at 0.5 millimolar ATP or 0.25 millimolar PPi. ATP-driven transport showed a distinct pH optimum at 7.3 while PPi-driven transport reached maximum rates at pH 7.8. Direct measurement of the H+- translocating enzyme activities revealed Km apparent values of 0.45 millimolar for ATPase (EC 3.6.1.3) and 23 micromolar for pyrophosphatase (PPase) (EC 3.6.1.1). In contrast to the coupled amino acid transport, ATPase and PPase activities had relative broad pH optima between 7 to 8 for ATPase and 8 to 9 for PPase. ATPase as well as ATP-driven transport was markedly inhibited by nitrate while PPase and PPi-coupled transport was not affected. The addition of ionophores inhibited phenylalanine transport suggesting the destruction of the electrochemical proton potential difference while the rate of ATP and PPi hydrolysis was stimulated. The uptake of other lipophilic amino acids like L-Trp, L-Leu, and L-Tyr was also stimulated by ATP. They seem to compete for the same carrier system. L-Ala, L-Val, D-Phe, and D-Leu did not influence phenylalanine transport suggesting a stereospecificity of the carrier system for L-amino acids having a relatively high hydrophobicity.
ISSN:0032-0889
1532-2548
DOI:10.1104/pp.89.4.1388