Parachloromercuribenzenesulfonic Acid : a potential tool for differential labeling of the sucrose transporter.

Vicia faba leaf discs without epidermis were pretreated with parachloromercuribenzenesulfonic acid (PCMBS), rinsed and incubated on [(14)C]sucrose (1 or 40 millimolar). Those sucrose concentrations were chosen as representative of the apparent uptake system 1 (1 millimolar) and system 2 (40 millimolar) previously characterized. Pretreatment with 0.5 millimolar PCMBS for 20 minutes inhibited system 1 and system 2 by about 70%.Addition of unlabeled sucrose during PCMBS-pretreatment protected the carrier(s) from the inhibition, whereas glucose, fructose, and sucrose analogs were unable to afford protection. At 1 millimolar [(14)C]sucrose, the protection resulted in a small but consistent reduction of normal inhibition (from 63 to 45%) for sucrose concentrations of 50 millimolar and more during pretreatment. Contrarily, at 40 millimolar [(14)C]sucrose, the protection increased linearly with the sucrose concentration in the pretreatment medium, and complete prevention of inhibition was reached for 250 millimolar sucrose.The protection was not due to exchange diffusion and was located in the veins. Michaelian kinetics indicated that PCMBS and sucrose compete with each other at the active site of the carrier.Among 14 compounds tested (sugars, amino-acids, hormones, (32)P), sucrose uptake was by far the most sensitive to PCMBS. Sucrose preferentially protected its carrier(s) from inhibition. Treatment with 20 millimolar cysteine or 20 millimolar dithioerythreitol reversed inhibition by PCMBS pretreatment.