The Study Of Functional Evolution Of Mitochondrial Carrier Family

Exploring metabolic evolution is a way to understand metabolic complexity. The substrate transport of mitochondrial carrier family (MCF) influences direct metabolic activities, making it possible to understand indirectly metabolic evolution from the view of functional evolution of MCF. However, the study of functional evolution of MCF does not mean that all the concrete structures of mitochondrial carriers (MCs) must first be gained. We treated MCF as a whole, divided MCF structure into transmembrane region (TR) and non-transmembrane region (NON-TR), and characterized their structural changes respectively by studying the evolutionary rules of the sequences within these two regions. Based on this, we studied the alternation of MCF structure and potential correlated functions of MCF.The data analysis indicates that the types of substrates transported by MCF were maintained during the period of metazoan evolution. However, the size of the substrates transported by MCs continuously diminished during the evolutionary process. We have found that the ratio of hydrophobic amino acids at specific helix-helix interfaces increases significantly during vertebrate evolution. Amino acid’s spatial positioning and the calculating of packing values both indicate the increase in the number of hydrophobic amino acids will lead to a more "tight" structure of the TR domain, which is in agreement with the diminishing size of substrates transported by vertebrate MCs. In addition, there is a significant increase in the number of carriers of MCF during the period of vertebrate evolution. Based on these observations, the paper postulates that the more "tight" TR structure generated by the increase of the hydrophobic amino acids at specific helix-helix interfaces enhances the substrate selectivity of MCF, reflecting the evolutionary trajectory of MCF during metazoan evolution.On the one hand, the strengthening of substrate selectivity means a kind of molecular adaptation; on the other hand, it will be embodied in H+ (the minimum sized substrate) transport. Then, it will facilitate a further study of molecular mechanism of transporting H+ for exploring metabolic complexity evolutionTransporting H+ from cytosol to mitochondrial matrix is typical of mitochondrial proton leakage. It has been proposed that the eukaryotic MCF located at the mitochondrial inner membrane may transport H+.However, it is difficult to demonstrate mitochondrial carriers-mediated proton translocation for the objects are all members of MCF rather than individual or specific carriers. The predicted functional glutamine at the c-terminal 10th site of Px(D/E)xx(K/R)xR motif of MCs (Q10) in eukaryotes could act as proton-buffer to transport H+, indicating that preventing the re-entering of matrix H+ is a requirement for H+ channel transport. The increased proportion of arginine at the 8th site of the even-numbered transmembrane helices in metazoans might trigger the transfer of more protons through the channel. Their re-entering would be prevented by the strengthened positively charged potential surface barrier involved by the increasing proportion of glutamine at the c-terminal 12th site of MCs motif (Q12).Q12 positively correlated with eukaryotes evolution (R=0.977, P<0.001) showed adaptation to the augmentative H+ transport in metazoan evolution. The simulation experiment of the augmentative H+ transport represented that the more H+ had a deleterious effect on fungi with their lower proportion of Q12. Together with the common sequence features responsible for transporting H+ in MCs, this provided evidence for the mechanism of MCs-mediated H+ transport. Then, an "improved proton-buffering model" was proposed for depicting the mechanism of MCs-mediated proton translocation. Based on this, it was proposed that the increased proportion of Q12 and R8 in metazoan evolution suggested the evolutionary mechanism of mitochondrial proton leakage.Based on the evolutionary, structural and biochemical evidences, this study not only reveals the evolutionary trajectory of MCF, but also provides insights into the evolutionary mechanism of proton leakage. This will provides some hints to the molecular mechanism of metabolic complexity.