Pussinen P J, Olkkonen V M, Jauhiainen M, Ehnholm C
Department of Biochemistry, National Public Health Institute, Helsinki, Finland.
J Lipid Res. 1997 Jul;38(7):1473-81.
Humans and the pig show marked similarities in lipoprotein metabolism; therefore, the pig has been used as a model in numerous nutritional studies. Pig plasma displays no activity of cholesteryl ester transfer protein (CETP), which is known to be responsible for half of the phospholipid mass transfer in human plasma, the other half being accounted for by the plasma phospholipid transfer protein (PLTP). This makes the pig an ideal subject for the study of PLTP structure and function. Here we report the molecular cloning of pig PLTP and the eukaryotic cell expression of its complementary DNA. Pig PLTP was found to share 93% amino acid sequence identity with human PLTP and 81% with mouse PLTP. Tissue expression of PLTP mRNA was examined by a method based on reverse transcription-polymerase chain reaction (RT-PCR) and solid-phase minisequencing in nine pig tissues. The highest PLTP mRNA levels were found in the pancreas, brain, lung, and liver. Medium from COS-1 cells expressing PLTP possessed phospholipid transfer activity, and the secreted recombinant PLTP was detectable by Western blotting in the culture supernatant. A mutant protein with a substitution of Cys at position 22 by Arg was found to display impaired secretion into growth medium indicating a role for cysteines in the correct folding of PLTP. This study forms the basis for future work on the structure-function relationships in pig PLTP.
人类和猪在脂蛋白代谢方面表现出显著的相似性；因此，猪已被用作众多营养研究中的模型。猪血浆中不存在胆固醇酯转移蛋白（CETP）的活性，已知该蛋白负责人类血浆中一半的磷脂质量转移，另一半则由血浆磷脂转移蛋白（PLTP）负责。这使得猪成为研究PLTP结构和功能的理想对象。在此，我们报告猪PLTP的分子克隆及其互补DNA的真核细胞表达。发现猪PLTP与人类PLTP的氨基酸序列同一性为93%，与小鼠PLTP的同一性为81%。通过基于逆转录-聚合酶链反应（RT-PCR）和固相微测序的方法，检测了9种猪组织中PLTP mRNA的组织表达情况。在胰腺、脑、肺和肝脏中发现PLTP mRNA水平最高。表达PLTP的COS-1细胞培养基具有磷脂转移活性，并且在培养上清液中通过蛋白质印迹法可检测到分泌的重组PLTP。发现一种在第22位的半胱氨酸被精氨酸取代的突变蛋白向生长培养基中的分泌受损，这表明半胱氨酸在PLTP的正确折叠中起作用。本研究为未来关于猪PLTP结构-功能关系的研究奠定了基础。
