Kim Kyung Hyun, Jia Baolei, Jeon Che Ok
Department of Life Science, Chung-Ang University, Seoul, South Korea.
Front Microbiol. 2017 Oct 20;8:2054. doi: 10.3389/fmicb.2017.02054. eCollection 2017.
, a moderately halophilic bacterium, accumulates a variety of compatible solutes including glycine betaine, glutamate, glutamine, proline, and ectoine to cope with osmotic stress. Non-targeted analysis of intracellular organic compounds using H-NMR showed that a large amount of trans-4-hydroxy-L-proline (Hyp), which has not been reported as a compatible solute in , was accumulated in response to high NaCl salinity, suggesting that Hyp may be an important compatible solute in . Candidate genes encoding proline 4-hydroxylase (PH-4), which hydroxylates L-proline to generate Hyp, were retrieved from the genome of through domain searches based on the sequences of known PH-4 proteins. A gene, HBHAL_RS11735, which was annotated as a multidrug DMT transporter permease in GenBank, was identified as the PH-4 gene through protein expression analysis in . The PH-4 gene constituted a transcriptional unit with a promoter and a rho-independent terminator, and it was distantly located from the proline biosynthetic gene cluster ( operon). Transcriptional analysis showed that PH-4 gene expression was NaCl concentration-dependent, and was specifically induced by chloride anion, similar to the operon. Accumulation of intracellular Hyp was also observed in other bacteria, suggesting that Hyp may be a widespread compatible solute in halophilic and halotolerant bacteria.
一种中度嗜盐细菌积累多种相容性溶质，包括甘氨酸甜菜碱、谷氨酸、谷氨酰胺、脯氨酸和四氢嘧啶，以应对渗透胁迫。使用氢核磁共振对细胞内有机化合物进行非靶向分析表明，在高氯化钠盐度下，大量反式-4-羟基-L-脯氨酸（Hyp）被积累，而Hyp在该细菌中尚未被报道为一种相容性溶质，这表明Hyp可能是该细菌中一种重要的相容性溶质。通过基于已知脯氨酸4-羟化酶（PH-4）蛋白序列的结构域搜索，从该细菌的基因组中检索到编码将L-脯氨酸羟基化生成Hyp的脯氨酸4-羟化酶（PH-4）的候选基因。一个在GenBank中被注释为多药DMT转运通透酶的基因HBHAL_RS11735，通过在该细菌中的蛋白质表达分析被鉴定为PH-4基因。PH-4基因与一个启动子和一个不依赖ρ因子的终止子构成一个转录单元，并且它与脯氨酸生物合成基因簇（操纵子）距离较远。转录分析表明，PH-4基因的表达依赖于氯化钠浓度，并且与操纵子一样，受到氯离子特异性诱导。在其他细菌中也观察到细胞内Hyp的积累，这表明Hyp可能是嗜盐和耐盐细菌中广泛存在的一种相容性溶质。
