Li Mohui, Yang Ziqing, Chen Sihua, Liu Zilu, Tong Li, Zheng Shaokui, Yang Dong
Gene Engineering and Biotechnology Beijing Key Laboratory, College of Life Sciences, Beijing Normal University, Beijing, 100875, China.
School of Environment, MOE Key Laboratory of Water and Sediment Sciences/State Key Lab of Water Environment Simulation, Beijing Normal University, Beijing 100875, China.
Int J Biol Macromol. 2024 Jan;254(Pt 3):128112. doi: 10.1016/j.ijbiomac.2023.128112. Epub 2023 Nov 14.
Bacterial hemoglobins play important roles inside the cell. Phylogenetically, they belong to three different families: the single domain hemoglobin, flavohemoglobin and truncated hemoglobin. Vitreoscilla hemoglobin (VHb) is the first characterized bacterial hemoglobin, and belongs to the single domain hemoglobin family. Heterologous expression of VHb promotes the growth of host cells under microaerobic conditions, and enhances the yield of products during fermentation. Although VHb has been widely applied in the biotechnology field, other bacterial hemoglobins have not demonstrated similar applications. In this study, we identified four bacterial hemoglobins from the microaerobic growing bacterium Sphaerotilus natans, including one flavohemoglobins (FHB) and three truncated hemoglobins (THB1, THB2 and THB3). Absorption spectrum studies validate the existent of the Soret peak and Q-band characteristic to heme and suggest heme groups in FHB and THB1 are hexa- or penta-coordinated, respectively. Our studies demonstrate that FHB and all three truncated hemoglobins have NADH oxidation and radical production activities, which is surprising since truncated hemoglobins do not have a reductase domain that could bind NADH. However, the M. tuberculosis HbN does not show these activities, indicating they are not universal among truncated hemoglobins. Docking studies suggest the nicotinamide ring of NADH may bind to the distal heme pocket of THB1, suggesting the direct electron transfer from NADH to heme might be possible. Our truncated hemoglobins also show peroxidase activities that in THB2 and THB3 could be inhibited by FdR, indicating possible interactions between FdR and truncate hemoglobins. Expression of FHB and THB1 in E. coli could promote cell growth. THB1 also enhances the production of limonene in an engineered E. coli strain, while VHb does not have this effect, which suggests that studies on truncated hemoglobins may lead to the discovery of new and more powerful tools that could have profound impact on biotechnology.
细菌血红蛋白在细胞内发挥着重要作用。从系统发育角度来看，它们属于三个不同的家族：单结构域血红蛋白、黄素血红蛋白和截短血红蛋白。透明颤菌血红蛋白（VHb）是首个被鉴定的细菌血红蛋白，属于单结构域血红蛋白家族。VHb的异源表达可促进宿主细胞在微需氧条件下的生长，并提高发酵过程中产物的产量。尽管VHb已在生物技术领域得到广泛应用，但其他细菌血红蛋白尚未展现出类似的应用。在本研究中，我们从微需氧生长的浮游球衣菌中鉴定出四种细菌血红蛋白，包括一种黄素血红蛋白（FHB）和三种截短血红蛋白（THB1、THB2和THB3）。吸收光谱研究证实了FHB和THB1中存在血红素特有的Soret峰和Q带特征，并表明FHB和THB1中的血红素基团分别为六配位或五配位。我们的研究表明，FHB和所有三种截短血红蛋白都具有NADH氧化和自由基产生活性，这令人惊讶，因为截短血红蛋白没有可结合NADH的还原酶结构域。然而，结核分枝杆菌HbN不显示这些活性，表明它们在截短血红蛋白中并不普遍存在。对接研究表明，NADH的烟酰胺环可能与THB1的远端血红素口袋结合，这表明NADH可能直接向血红素进行电子转移。我们的截短血红蛋白还表现出过氧化物酶活性，其中THB2和THB3中的过氧化物酶活性可被FdR抑制，这表明FdR与截短血红蛋白之间可能存在相互作用。FHB和THB1在大肠杆菌中的表达可促进细胞生长。THB1还可提高工程化大肠杆菌菌株中柠檬烯的产量，而VHb则没有这种效果，这表明对截短血红蛋白的研究可能会发现新的、更强大的工具，这些工具可能会对生物技术产生深远影响。
