Date Published: June 8, 2018
Publisher: Public Library of Science
Author(s): Mohammad Uzzal Hossain, Taimur Md. Omar, Iftekhar Alam, Keshob Chandra Das, A. K. M. Mohiuddin, Chaman Ara Keya, Md. Salimullah, Patrick Jon Biggs.
The bacterial species Campylobacter jejuni RM1221 (CjR) is the primary cause of campylobacteriosis which poses a global threat for human health. Over the years the efficacy of antibiotic treatment is becoming more fruitless due to the development of multiple drug resistant strains. Therefore, identification of new drug targets is a valuable tool for the development of new treatments for affected patients and can be obtained by targeting essential protein(s) of CjR. We conducted this in silico study in order to identify therapeutic targets by subtractive CjR proteome analysis. The most important proteins of the CjR proteome, which includes chokepoint enzymes, plasmid, virulence and antibiotic resistant proteins were annotated and subjected to subtractive analyses to filter out the CjR essential proteins from duplicate or human homologous proteins. Through the subtractive and characterization analysis we have identified 38 eligible therapeutic targets including 1 potential vaccine target. Also, 12 potential targets were found in interactive network, 5 targets to be dealt with FDA approved drugs and one pathway as potential pathway based drug target. In addition, a comprehensive database ‘CampyNIBase’ has also been developed. Besides the results of this study, the database is enriched with other information such as 3D models of the identified targets, experimental structures and Expressed Sequence Tag (EST) sequences. This study, including the database might be exploited for future research and the identification of effective therapeutics against campylobacteriosis. URL: (http://nib.portal.gov.bd/site/page/4516e965-8935-4129-8c3f-df95e754c562#Banner).
The genus Campylobacter is composed of a wide variety of non-spore forming Gram-negative bacteria that are predominantly rod or spiral shaped . Most Campylobacter infections are acquired through contaminated food and two species, Campylobacter coli and Campylobacter jejuni are the primary cause of the human disease termed as campylobacteriosis [2,3]. Campylobacter jejuni (C. jejuni) is the species that induces acute gastroenteritis and bacterial food poisoning in infected patients. Normal infection with C. jejuni causes uncomplicated gastroenteritis but severe infection may result in abdominal cramps, fever or even serious diseases like diarrhea, GuillainBarré Syndrome or Miller Fischer Syndrome [4–6]. C. jejuni infection is acquired via numerous sources associated with lack of awareness such as undercooked livestock meat, poultry, unpasteurized milk or contaminated water sources .
The identification of drug targets was carried out in three phases. In phase-I, major proteins of CjR genome were collected from different sources. These proteins include chokepoint enzymes, plasmid proteins, virulent proteins and antibiotic resistant proteins. In phase-II, subtractive analyses were carried out through different steps by excluding human homologues proteins and collecting important proteins required for the survival of CjR. In phase-III, potential drug targets found from subtractive analyses in phase-II are characterized. The entire work flow can be seen as a flowchart (Fig 1).
To our knowledge the current subtractive analysis performed in this study is the first report on computational analysis to identify and characterize therapeutic targets of CjR. We have selected CjR strainof campylobacter which is most prevalent in Bangladesh among the others. In molecular level, CjR strain is reported as one of the most frequent etiologic agent to cause about 65% diarrheal diseases in different clinical isolates in Bangladesh [74,75]. Besides this, technically to work with CjR strain in Bioinformatics analysis is advantageous asthis strain is well annotated with complete genome in KEGG and KAAS database and other bioinformatics resources as well. We have exploited total proteins of CjR for pathway based drug design. Whole proteome annotation identified a substantial number (2218) of protein sequences through various databases and literature searches. In this study, we have particularly focused on chokepoint enzymes, plasmid proteins, antibiotic resistance genes which are essential for CjR. In a metabolic reaction, only a specific substrate can be consumed and specific product produced during catalysis by a specific enzyme. This type of reaction is known as a chokepoint reaction and the enzyme that catalyzes the reaction is called a chokepoint enzyme. Therefore, certain chokepoint enzymes of a bacterial strain have potential to be a promising drug target . As plasmid proteins can confer unique characteristics to a specific strain, we have collected their sequences through literature survey. In addition, antibiotic resistances by C. jejuni are mutations in certain genes. C. jejuni strains that have acquired multiple antibiotic resistances (MAR) overexpress cmeB compared to normal strains . Many C. jejuni strains harbour point mutations in the gyrA gene to facilitate their resistance to fluoroquinolone . Moreover, another resistance gene called aphA-7, which encodes a kanamycin resistant phosphotransferase, is found to be native in the genome of numerous Campylobacter species . Usually, successful pathogenic bacteria rely on multiple virulent factors for survival and effective replication inside a host which in turn makes them potential drug targets (S1–S5 Tables).
The approach used in this study could corroborate to a powerful channel of analysis with rational accuracy toward identification of important essential genes in bacteria. This study identified one vaccine, 2 drug targets and 1 pathway based drug target. These potential therapeutic targets could be further validated experimentally through the drug and vaccine design pipelines. Together with 3D structures and other data, a comprehensive database ‘CampyNIBase’ has been developed to assist future CjR research.