Dataset Information

Profiling of cow urinary proteins using various extraction methods reveals more than 1550 proteins

ABSTRACT: Urine can help in diagnosis of different diseases including cancer and other patho-physiological conditions. Urine proteome studies have been mainly human- centric. No information is available on urinary proteome from bovine till date. In the present study, we have used 3 protein extraction methods such as ammonium sulphate precipitation, ProteoSpin column and diafiltration method from bovine urine for identification of urinary proteome. The tryptic peptides generated after In-gel and In-solution method were identified using LC/MS/MS (ESI-qTOF) which resulted in identification of 1582 proteins. In-gel trypsin digestion method revealed more protein (1191) in comparison to in-solution digestion method (541). Maximum proteins were identified in ammonium sulphate precipitation method (938) followed by ProteoSpin (606) and diafiltration (444) methods respectively. The profile of the identified proteins were compared with human urinary proteome of which 311 bovine urinary proteins matched with human. An exclusive list of 38 bovine urinary proteins with high protein scores were listed which are absent in human urine. All identified proteins were analyzed according to Gene Ontology which were classified according to cellular component, biological processes and molecular functions. This study reports for the first time an exclusive evidence of more than 1550 proteins in urine of healthy cow donors.

INSTRUMENT(S): maXis

ORGANISM(S): Bos Taurus (bovine)

TISSUE(S): Urine

SUBMITTER: Ashok Kumar Mohanty

LAB HEAD: Ashok Kumar Mohanty

PROVIDER: PXD002038 | Pride | 2023-03-22

REPOSITORIES: Pride

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Publications

Profiling of urinary proteins in Karan Fries cows reveals more than 1550 proteins.

Bathla Shveta S Rawat Preeti P Baithalu Rubina R Yadav Munna Lal ML Naru Jasmine J Tiwari Anurag A Kumar Sudarshan S Balhara Ashok K AK Singh Surender S Chaudhary Suman S Kumar Rajesh R Lotfan Masoud M Behare Pradip P Phulia Sushil K SK Mohanty Tushar K TK Kaushik Jai K JK Nallapeta Shivramaiah S Singh Inderjeet I Ambatipudi Srinivas K SK Mohanty Ashok K AK

Journal of proteomics 20150602 Pt A

Urine is a non-invasive source of biological fluid, which reflects the physiological status of the mammals. We have profiled the cow urinary proteome and analyzed its functional significance. The urine collected from three healthy cows was concentrated by diafiltration (DF) followed by protein extraction using three methods, namely methanol, acetone, and ammonium sulphate (AS) precipitation and Proteo Spin urine concentration kit (PS). The quality of the protein was assessed by two-dimensional g ...[more]

PMID: 26021477

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Project description:Pseudomonas aeruginosa is one of the most frequent pathogen dominant in complicated urinary tract infections (UTI). To unravel the adaptation strategies of P. aeruginosa to the conditions in the urinary tract and to define the underlying regulatory network an artificial growth system mimicking the conditions in the urinary tract was established. Transcriptome analyses were used to investigate the physiological status of P. aeruginosa under this conditions. We performed comparisons to identify genes induced under artificial urinary tract conditions to unravel the adaptive strategies and the underlying regulatory network used by Pseudomonas aeruginosa during urinary tract infections using Affimetrix GeneChips. Pseudomonas aeruginosa wild type strain PAO1 was grown in an artificial in vitro growth system mimicking the conditions in the urinary tract. Therefore, biofilms were grown on the surface of membrane filters placed on agar plates at 37 Â°C up to the late logarithmic state under aerobic and anaerobic conditions (incubated in an anaerobic beanch). An artificial urine medium (AUM) simulating the averaged urine of an human adult was used as nutrient souce. 10-fold diluted Luria Bertani (LB)-medium was used as reference medium. For growth under oxygen depletion the media were supplemented with 50 mM KNO3 to sustain anaerobic respiration. The biofilms were harveted at this time points and resuspsended in 0.9% (w/v) NaCl. The OD578 of biofilm suspension was 0.8 for all tested conditions. First comparison: Identification of genes induced or repressed under aerobic conditions in the P. aeruginosa wild type PAO1. Here we compared the transcriptome profile of P. aeruginosa PAO1 grown aerobically for 18 h to the late logarithmic phase in biofilms on AUM with the transcriptome profile of the PAO1 strain, which was grown aerobically for 18 h to the late logarithmic phase in biofilms on 10-fold diluted LB. Second comparison: Identification of genes induced or repressed under anaerobic conditions in the P. aeruginosa wild type PAO1. Here we compared the transcriptome profile of P. aeruginosa PAO1 grown anaerobically for 2 days up to the late logarithmic phase in biofilms on AUM supplemented with 50 mM nitrate with the transcriptome profile of the PAO1 strain, which was grown anaerobically for 2 days up to the late logarithmic phase in biofilms on 10-fold diluted LB supplemented with 50 mM nitrate.

Project description:Long term-catheterization leads inevitably to a catheter-associated bacteriuria caused by multispecies bacterial biofilms growing on and in the catheters. The overall goal of the presented study was (I) to unravel bacterial community structure and function of such an uropathogenic biofilm and (II) to elucidate the interplay between bacterial virulence and the human immune system within the urine. To this end, a metaproteomics approach combined with in vitro proteomics analyses was employed to investigate both, the pro- and eukaryotic protein inventory. Our proteome analyses demonstrated that the biofilm of the investigated catheter is dominated by three bacterial species, i.e. Pseudomonas aeruginosa, Morganella morganii and Bacteroides sp., and identified iron limitation as one of the major challenges in the bladder environment. In vitro proteome analysis of P. aeruginosa and M. morganii isolated from the biofilm revealed that those opportunistic pathogens are able to overcome iron restriction via the production of siderophores and high expression of corresponding receptors. Notably, a comparison of in vivo and in vitro protein profiles of P. aeruginosa and M. morganii also indicated that the bacteria employ different strategies to adapt to the urinary tract. Whilst P. aeruginosa seems to express secreted and surface-exposed proteases to escape the human innate immune system and metabolizes amino acids, M. morganii is able to take up sugars and to degrade urea. Most interestingly, a comparison of urine protein profiles of three long-term catheterized patients and three healthy control persons demonstrated the elevated level of proteins associated to neutrophils, macrophages and complement system in the patient urine, which might point to a specific activation of the innate immune system in response to biofilm-associated urinary tract infections. We thus hypothesize that the often asymptomatic nature of CAUTI´s might be based on a fine-tuned balance between the expression of bacterial virulence factors and the human immune system. Long term-catheterization leads inevitably to a catheter-associated bacteriuria caused by multispecies bacterial biofilms growing on and in the catheters. The overall goal of the presented study was (I) to unravel bacterial community structure and function of such an uropathogenic biofilm and (II) to elucidate the interplay between bacterial virulence and the human immune system within the urine. To this end, a metaproteomics approach combined with in vitro proteomics analyses was employed to investigate both, the pro- and eukaryotic protein inventory. Our proteome analyses demonstrated that the biofilm of the investigated catheter is dominated by three bacterial species, i.e. Pseudomonas aeruginosa, Morganella morganii and Bacteroides sp., and identified iron limitation as one of the major challenges in the bladder environment. In vitro proteome analysis of P. aeruginosa and M. morganii isolated from the biofilm revealed that those opportunistic pathogens are able to overcome iron restriction via the production of siderophores and high expression of corresponding receptors. Notably, a comparison of in vivo and in vitro protein profiles of P. aeruginosa and M. morganii also indicated that the bacteria employ different strategies to adapt to the urinary tract. Whilst P. aeruginosa seems to express secreted and surface-exposed proteases to escape the human innate immune system and metabolizes amino acids, M. morganii is able to take up sugars and to degrade urea. Most interestingly, a comparison of urine protein profiles of three long-term catheterized patients and three healthy control persons demonstrated the elevated level of proteins associated to neutrophils, macrophages and complement system in the patient urine, which might point to a specific activation of the innate immune system in response to biofilm-associated urinary tract infections. We thus hypothesize that the often asymptomatic nature of CAUTI´s might be based on a fine-tuned balance between the expression of bacterial virulence factors and the human immune system.

Dataset Information

Profiling of cow urinary proteins using various extraction methods reveals more than 1550 proteins

Publications

Profiling of urinary proteins in Karan Fries cows reveals more than 1550 proteins.

Similar Datasets

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