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首页> 外文期刊>Journal of proteome research >Sub-speciating Campylobacter jejuni by proteomic analysis of its protein biomarkers and their post-translational modifications
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Sub-speciating Campylobacter jejuni by proteomic analysis of its protein biomarkers and their post-translational modifications

机译：通过蛋白质组学蛋白质组标记及其翻译后修饰对空肠弯曲菌进行分类

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摘要

We have identified several protein biomarkers of three Campylobacter jejuni strains (RM1221, RM1859, and RM3782) by proteomic techniques. The protein biomarkers identified are prominently observed in the time-of-flight mass spectra (TOF MS) of bacterial cell lysate supernatants ionized by matrix-assisted laser desorption/ionization (MALDI). The protein biomarkers identified were: DNA-binding protein HU, translation initiation factor IF-1, cytochrome c 553, a transthyretin-like periplasmic protein, chaperonin GroES, thioredoxin Trx, and ribosomal proteins: L7/L12 (50S), L24 (50S), S16 (30S), L29 (50S), and S15 (30S), and conserved proteins similar to strain NCTC 11168 proteins Cj1164 and Cj1225. The protein biomarkers identified appear to represent high copy, intact proteins. The significant findings are as follows: (1) Biomarker mass shifts between these strains were due to amino acid substitutions of the primary polypeptide sequence and not due to changes in post-translational modifications (PTMs). (2) If present, a PTM of a protein biomarker appeared consistently for all three strains, which supported that the biomarker mass shifts observed between strains were not due to PTM variability. (3) The PTMs observed included N-terminal methionine (N-Met) cleavage as well as a number of other PTMs. (4) It was discovered that protein biomarkers of C. jejuni (as well as other thermophilic Campylobacters) appear to violate the N-Met cleavage rule of bacterial proteins, which predicts N-Met cleavage if the penultimate residue is threonine. Two protein biomarkers (HU and 30S ribosomal protein S16) that have a penultimate threonine residue do not show N-Met cleavage. In all other cases, the rule correctly predicted N-Met cleavage among the biomarkers analyzed. This exception to the N-Met cleavage rule has implications for the development of bioinformatics algorithms for protein/pathogen identification. (5) There were fewer biomarker mass shifts between strains RM1221 and RM1859 compared to strain RM3782. As the mass shifts were due to the frequency of amino acid substitutions (and thus underlying genetic variations), this suggested that strains RM1221 and RM1859 were phylogenetically closer to one another than to strain RM3782 (in addition, a protein biomarker prominent in the spectra of RM1221 and RM1859 was absent from the RM3782 spectrum due to a nonsense mutation in the gene of the biomarker). These observations were confirmed by a nitrate reduction test, which showed that RM1221 and RM1859 were C. jejuni subsp. jejuni whereas RM3782 was C. jejuni subsp. doylei. This result suggests that detection/identification of protein biomarkers by pattern recognition and/or bioinformatics algorithms may easily subspeciate bacterial microorganisms. (6) Finally, the number and variation of PTMs detected in this relatively small number of protein biomarkers suggest that bioinformatics algorithms for pathogen identification may need to incorporate many more possible PTMs than suggested previously in the literature.
机译：我们已经通过蛋白质组学技术鉴定了三种空肠弯曲杆菌菌株（RM1221，RM1859和RM3782）的几种蛋白质生物标记。在通过基质辅助激光解吸/电离（MALDI）电离的细菌细胞裂解液上清液的飞行时间质谱图（TOF MS）中，观察到的蛋白质生物标记物得到了显着观察。鉴定出的蛋白质生物标志物是：DNA结合蛋白HU，翻译起始因子IF-1，细胞色素c 553，甲状腺素蛋白样周质蛋白，伴侣蛋白GroES，硫氧还蛋白Trx和核糖体蛋白：L7 / L12（50S），L24（50S ），S16（30S），L29（50S）和S15（30S）以及与菌株NCTC 11168蛋白Cj1164和Cj1225类似的保守蛋白。鉴定出的蛋白质生物标志物似乎代表了高拷贝的完整蛋白质。重大发现如下：（1）这些菌株之间的生物标志物质量转移是由于一级多肽序列的氨基酸取代，而不是由于翻译后修饰（PTM）的改变。 （2）如果存在，则所有三个菌株的蛋白质生物标志物的PTM均一致出现，这支持在菌株之间观察到的生物标志物质量变化并非归因于PTM变异性。 （3）观察到的PTM包括N端甲硫氨酸（N-Met）裂解以及许多其他PTM。 （4）发现空肠弯曲杆菌的蛋白质生物标志物（以及其他嗜热弯曲杆菌）似乎违反了细菌蛋白质的N-Met裂解规则，如果倒数第二个残基是苏氨酸，则可以预测N-Met裂解。具有倒数第二个苏氨酸残基的两个蛋白质生物标记（HU和30S核糖体蛋白S16）没有显示N-Met裂解。在所有其他情况下，该规则正确预测了所分析生物标志物中的N-Met裂解。 N-Met裂解规则的这一例外情况对蛋白质/病原体鉴定的生物信息学算法的发展产生了影响。 （5）与菌株RM3782相比，菌株RM1221和RM1859之间的生物标志物质量转移较少。由于质量转移是由于氨基酸置换的频率（以及潜在的遗传变异）引起的，因此表明菌株RM1221和RM1859在系统发育上比菌株RM3782更接近彼此（此外，在蛋白质谱中的蛋白质生物标记很显着）。由于生物标记基因的无义突变，RM3782谱中没有RM1221和RM1859。这些观察结果通过硝酸盐还原测试得到证实，该测试表明RM1221和RM1859是空肠弯曲杆菌亚种。空肠，而RM3782是空肠弯曲杆菌亚种。多雷该结果表明通过模式识别和/或生物信息学算法对蛋白质生物标志物的检测/鉴定可能容易地将细菌微生物亚种化。 （6）最后，在这种相对较少的蛋白质生物标记物中检测到的PTM的数量和变异表明，用于病原体鉴定的生物信息学算法可能需要比文献中先前建议的方法纳入更多的可能PTM。

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