Kinetics and substrate specificity of membrane-reconstituted peptide transporter DtpT of Lactococcus lactis
Kinetics and substrate specificity of membrane-reconstituted peptide transporter DtpT of Lactococcus lactis
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DOI：
10.1128/jb.182.9.2530-2535.2000
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发表时间：
2000-05-01
影响因子：
3.2
通讯作者：
Poolman, B
中科院分区：
文献类型：
--
作者：
Fang, G;Konings, WN;Poolman, B
将乳酸乳球菌的多肽转运蛋白DtpT纯化重组为洗涤剂不稳定脂质体，利用Pro-[C-14]Ala作为报告肽，在多种多肽或多肽模拟物存在的情况下，测定了该转运蛋白在蛋白脂质体系统中的动力学和底物特异性。DtpT蛋白似乎对二肽和三肽具有特异性，对具有至少一个疏水残基的肽具有最高的亲和力。氨基酸的疏水性、大小或电荷对二肽氨基端和羧基端位置的影响是不同的。游离氨基酸，ω -氨基脂肪酸化合物，或肽超过三个氨基酸残基不与DtpT相互作用。对于与DtpT的高亲和力相互作用，肽需要具有游离的氨基和羧基端，L构型的氨基酸和反肽键。将DtpT与真核生物同源物PepT(1)和PepT(2)的特异性进行比较，发现细菌转运体对底物的识别更具限制性。
The peptide transport protein DtpT of Lactococcus lactis was purified and reconstituted into detergent-destabilized liposomes, The kinetics and substrate specificity of the transporter in the proteoliposomal system were determined, using Pro-[C-14]Ala as a reporter peptide in the presence of various peptides or peptide mimetics. The DtpT protein appears to be specific for di- and tripeptides, with the highest affinities for peptides with at least one hydrophobic residue. The effect of the hydrophobicity, size, or charge of the amino acid was different for the amino- and carboxyl-terminal positions of dipeptides. Free amino acids, omega-amino fatty acid compounds, or peptides with more than three amino acid residues do not interact with DtpT. For high-affinity interaction with DtpT, the peptides need to have free amino and carboxyl termini, amino acids in the L configuration, and trans-peptide bonds. Comparison of the specificity of DtpT with that of the eukalyotic homologues PepT(1) and PepT(2) shows that the bacterial transporter is more restrictive in its substrate recognition.