The permeabilized SecY protein-translocation channel can serve as a nonspecific sugar transporter

Sen Mei; Chong Xie; Hao Mi; Chuang Xue; Qiang Guo; Guang-Qing Du; Guo-Bei Li; Cai-Xia Li; Ya-Nan Qu; Ming-Hao Xiong; Yang Jiang; Tian-Wei Tan; Shang-Tian Yang; Li-Hai Fan

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/378786 doi: bioRxiv preprint 9 substitutions at Ile 191 and Ile 408 (SecY (ΔP, IIIGIG)) provoked a 1.27-fold increase in 171 permeation of 2-deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl)amino]-D-glucose (2-172 NBDG) which has almost the same V A and D AB as lactose (Appendix Table S1 ). By co-173 expression of SecY (ΔP, IIIGIG) and SCVE, E. coli (ΔLacY) exhibited a moderate 174 growth on lactose 50 h after cell inoculation (Fig 4A) . 175 The benefits of using the SecY channel mutant for sugar transport 176 Unlike free diffusion which is linear in the concentration difference, the rate of active 177 transport or facilitated diffusion tends to approach saturation as the external sugar content The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/378786 doi: bioRxiv preprint 11 Shanghai Institutes for Biological Sciences (CAS), respectively. P thl is the thlA (cac2873) 217 promoter region of C. acetobutylicum ATCC 824. Luria-Bertani (LB) medium contains: 218 10 g/L tryptone, 5 g/L yeast extract, and 10 g/L NaCl. M9 mineral medium (without 219 sugars) contains: 7.52 g/L Na 2 HPO 4 ·2H 2 O, 3 g/L KH 2 PO 4 , 0.5 g/L NaCl, 0.5 g/L NH 4 Cl, (Datsenko & Wanner, 2000) . Briefly, PCR product was generated by using 237 primers (Appendix Table S2 ) that are homologous to regions adjacent to the gene to be 238 inactivated and template pKD13 carrying kanamycin resistance gene that is flanked by 239 All rights reserved. No reuse allowed without permission.