Department of Biochemistry and Molecular Biology, Dalhousie University, Halifax, Nova Scotia, Canada.
PLoS One. 2013 Jul 30;8(7):e70396. doi: 10.1371/journal.pone.0070396. Print 2013.
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key enzyme of the glycolytic pathway, reversibly catalyzing the sixth step of glycolysis and concurrently reducing the coenzyme NAD(+) to NADH. In photosynthetic organisms a GAPDH paralog (Gap2 in Cyanobacteria, GapA in most photosynthetic eukaryotes) functions in the Calvin cycle, performing the reverse of the glycolytic reaction and using the coenzyme NADPH preferentially. In a number of photosynthetic eukaryotes that acquired their plastid by the secondary endosymbiosis of a eukaryotic red alga (Alveolates, haptophytes, cryptomonads and stramenopiles) GapA has been apparently replaced with a paralog of the host's own cytosolic GAPDH (GapC1). Plastid GapC1 and GapA therefore represent two independent cases of functional divergence and adaptations to the Calvin cycle entailing a shift in subcellular targeting and a shift in binding preference from NAD(+) to NADPH.
We used the programs FunDi, GroupSim, and Difference Evolutionary-Trace to detect sites involved in the functional divergence of these two groups of GAPDH sequences and to identify potential cases of convergent evolution in the Calvin-cycle adapted GapA and GapC1 families. Sites identified as being functionally divergent by all or some of these programs were then investigated with respect to their possible roles in the structure and function of both glycolytic and plastid-targeted GAPDH isoforms.
In this work we found substantial evidence for convergent evolution in GapA/B and GapC1. In many cases sites in GAPDHs of these groups converged on identical amino acid residues in specific positions of the protein known to play a role in the function and regulation of plastid-functioning enzymes relative to their cytosolic counterparts. In addition, we demonstrate that bioinformatic software like FunDi are important tools for the generation of meaningful biological hypotheses that can then be tested with direct experimental techniques.
甘油醛-3-磷酸脱氢酶（GAPDH）是糖酵解途径的关键酶，可逆地催化糖酵解的第六步反应，同时将辅酶 NAD（+）还原为 NADH。在光合生物中，GAPDH 同工酶（蓝细菌中的 Gap2，大多数光合真核生物中的 GapA）在卡尔文循环中发挥作用，进行糖酵解的逆反应，并优先使用辅酶 NADPH。在一些通过真核红藻的二次内共生获得质体的光合真核生物中（涡鞭毛藻、甲藻、隐藻和不等鞭毛类），GapA 显然被宿主自身细胞质 GAPDH 的同工酶（GapC1）所取代。因此，质体 GapC1 和 GapA 代表了两种独立的功能分化和对卡尔文循环的适应案例，涉及亚细胞靶向的转变以及从 NAD（+）到 NADPH 的结合偏好的转变。
我们使用 FunDi、GroupSim 和 Difference Evolutionary-Trace 程序来检测这两组 GAPDH 序列功能分化涉及的位点，并鉴定卡尔文循环适应的 GapA 和 GapC1 家族中潜在的趋同进化案例。通过所有或部分这些程序鉴定为功能分化的位点，然后研究它们在糖酵解和质体靶向 GAPDH 同工型的结构和功能中的可能作用。
在这项工作中，我们发现了 GapA/B 和 GapC1 趋同进化的大量证据。在许多情况下，这些组的 GAPDH 中的位点在蛋白质的特定位置收敛到相同的氨基酸残基，这些位置已知在质体功能酶的功能和调节中发挥作用，相对于它们的细胞质对应物。此外，我们证明了像 FunDi 这样的生物信息学软件是生成有意义的生物学假设的重要工具，然后可以用直接的实验技术来验证这些假设。
