Abstract
BackgroundPlant sucrose uptake transporters (SUTs) are H+/sucrose symporters related to the major facilitator superfamily (MFS). SUTs are essential for plant growth but little is known about their transport mechanism. Recent work identified several conserved, charged amino acids within transmembrane spans (TMS) in SUTs that are essential for transport activity. Here we further evaluated the role of one of these positions, R188 in the fourth TMS of OsSUT1, a type II SUT.ResultsThe OsSUT1(R188K) mutant, studied by expression in plants, yeast, and Xenopus oocytes, did not transport sucrose but showed a H+ leak that was blocked by sucrose. The H+ leak was also blocked by β-phenyl glucoside which is not translocated by OsSUT1. Replacing the corresponding Arg in type I and type III SUTs, AtSUC1(R163K) and LjSUT4(R169K), respectively, also resulted in loss of sucrose transport activity. Fluorination at the glucosyl 3 and 4 positions of α-phenyl glucoside greatly decreased transport by wild type OsSUT1 but did not affect the ability to block H+ leak in the R188K mutant.ConclusionOsSUT1 R188 appears to be essential for sucrose translocation but not for substrate interaction that blocks H+ leak. Therefore, we propose that an additional binding site functions in the initial recognition of substrates. The corresponding Arg in type I and III SUTs are equally important. We propose that R188 interacts with glucosyl 3-OH and 4-OH during translocation.
Highlights
Plant sucrose uptake transporters (SUTs) are H+/sucrose symporters related to the major facilitator superfamily (MFS)
The OsSUT1(R188K) mutant does not transport sucrose, yet the addition of sucrose induces a positive shift in current under voltage clamp conditions in oocytes expressing the mutant [19] (Figure 1B). β-phenyl glucoside is a substrate for Type I and III SUTs, but is not transported by Type II SUTs such as OsSUT1 [11,40]
R188 in OsSUT1 was identified as a charged amino acid within the fourth transmembrane spans (TMS) that is important for transport activity [19]
Summary
Plant sucrose uptake transporters (SUTs) are H+/sucrose symporters related to the major facilitator superfamily (MFS). SUTs are essential for plant growth but little is known about their transport mechanism. Sucrose transporters (SUTs or SUCs) are membrane proteins that facilitate the uptake of sucrose into the cytoplasm [2]. Driven by the electrochemical H+ gradient across the membrane, SUT proteins transport both sucrose and H+ into the cytoplasm at a ratio of 1:1 [3,4,5]. Type II transporters are present in all plants, and in monocots they are considered to function in phloem loading [9,12]. Each plant species has at least one Type III SUT, which is localized in the vacuolar membrane of cells [13,14,15]
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