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Background: Alterations at the molecular level in spermatozoa and seminal plasma can affect male fertility. The objective of this study was to determine if analysis of differential expression of proteins in varying semen parameters can serve as potential biomarkers for male infertility., Methods: The differential expression of proteins in the seminal plasma of men based on sperm count and morphology were examined utilizing proteomic tools. Subjects were categorized based on sperm concentration and morphology into 4 groups: 1) normal sperm count and normal morphology (NN); 2) normal sperm count and abnormal morphology (NA); 3) oligozoospermia and normal morphology (ON); and 4) oligozoospermia and abnormal morphology (OA). Proteomic analysis was performed by LC-MS/MS followed by functional bioinformatics analysis. Protein distribution in the NA, ON and OA groups was compared with that of the NN group., Results: Twenty proteins were differentially expressed among the 4 groups. Among the unique proteins identified, 3 were downregulated in the NA group, 1 in the ON group and 1 in the OA group while 2 were upregulated in the ON and OA groups. The functional analysis 1) identified biological regulation as the major processes affected and 2) determined that most of the identified proteins were of extracellular origin., Conclusions: We have identified proteins that are over-or underexpressed in the seminal plasma of men with poor sperm quality. The distinct presence of some of the proteins may serve as potential biomarkers and provide insight into the mechanistic role played by these proteins in male infertility. Further studies using Western Blot analysis are required to validate these findings.

Objective: To identify the relative abundance of proteins in pooled reactive oxygen species (ROS)-positive (ROS+) and ROS-negative (ROS-) semen samples with the use of two-dimensional differential in-gel electrophoresis (2D-DIGE)., Design: Spermatozoa suspensions from ROS+ and ROS- groups by 2D-DIGE analysis., Setting: Tertiary hospital., Patient(s): 20 donors and 32 infertile men., Intervention(s): Seminal ejaculates evaluated for semen and proteomic analysis., Main Outcome Measure(s): Semen samples from 20 donors and 32 infertile men were pooled, divided into ROS+ and ROS- groups based on the cutoff value of <20 relative light units/s/10(6) sperm and frozen. From each pooled group, spermatozoa were labeled with Cy3/Cy5 fluorescent dye. Duplicate 2D-DIGE gels were run. Image analysis was performed with the use of Decider software. Protein spots exhibiting ≥1.5-fold difference in intensity were excised from the preparatory gel and identified by liquid chromatography-mass spectrometry. Data were analyzed with the use of Sequest and Blast programs., Result(s): A total of 1,343 protein spots in gel 1 (ROS-) and 1,265 spots in gel 2 (ROS+) were detected. The majority of protein spots had similar expression, with 31 spots were differentially expressed. Six spots were significantly decreased and 25 increased in the ROS- sample compared with the ROS+ sample., Conclusion(s): Significantly different expression of protective proteins against oxidative stress was found in ROS-compared with ROS+ samples. These differences may explain the role of oxidation species in the pathology of male infertility., (Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

In this study, the sperm protein profile was compared between fertile and infertile men using 2-dimensional gel electrophoresis, liquid chromatography mass spectrometer analysis, and matrix-assisted laser desorption ionization-time-of-flight-mass spectrometry. Four unique proteins, semenogelin II precursor, prolactin-induced protein, clusterin isoform 1, and prostate-specific antigen isoform 1 preproprotein, were predominantly present in the semen of healthy men; however, semenogelin II precursor and clusterin isoform 1 were not seen in the semen of infertile men, suggesting unique differences in the spermatozoa protein profiles of fertile and infertile men., (Copyright © 2011 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Background: Male-factor infertility presents a vexing problem for many reproductively active couples. Many studies have focused on abnormal sperm parameters. Recent advances in proteomic techniques, especially in mass spectrometry, have aided in the study of sperm and more specifically, sperm proteins. The aim of this study was to review the current literature on the various proteomic techniques, and their usefulness in diagnosing sperm dysfunction and potential applications in the clinical setting., Methods: Review of PubMed database. Key words: spermatozoa, proteomics, protein, proteome, 2D-PAGE, mass spectrometry., Results: Recently employed proteomic methods, such as two-dimensional polyacrylamide gel electrophoresis, mass spectrometry, and differential in gel electrophoresis, have identified numerous sperm-specific proteins. They also have provided a further understanding of protein function involved in sperm processes and for the differentiation between normal and abnormal states. In addition, studies on the sperm proteome have demonstrated the importance of post-translational modifications, and their ability to bring about physiological changes in sperm function. No longer do researchers believe that in order for them to elucidate the biochemical functions of genes, mere knowledge of the human genome sequence is sufficient. Moreover, a greater understanding of the physiological function of every protein in the tissue-specific proteome is essential in order to unravel the biological display of the human genome., Conclusion: Recent advances in proteomic techniques have provided insight into sperm function and dysfunction. Several multidimensional separation techniques can be utilized to identify and characterize spermatozoa. Future developments in bioinformatics can further assist researchers in understanding the vast amount of data collected in proteomic studies. Moreover, such advances in proteomics may help to decipher metabolites which can act as biomarkers in the detection of sperm impairments and to potentially develop treatment for infertile couples.Further comprehensive studies on sperm-specific proteome, mechanisms of protein function and its proteolytic regulation, biomarkers and functional pathways, such as oxidative-stress induced mechanisms, will provide better insight into physiological functions of the spermatozoa. Large-scale proteomic studies using purified protein assays will eventually lead to the development of novel biomarkers that may allow for detection of disease states, genetic abnormalities, and risk factors for male infertility. Ultimately, these biomarkers will allow for a better diagnosis of sperm dysfunction and aid in drug development.

Background: Seminal plasma serves as a natural reservoir of antioxidants. It helps to remove excessive formation of reactive oxygen species (ROS) and consequently, reduce oxidative stress. Proteomic profiling of seminal plasma proteins is important to understand the molecular mechanisms underlying oxidative stress and sperm dysfunction in infertile men. Methods: This prospective study consisted of 52 subjects: 32 infertile men and 20 healthy donors. Once semen and oxidative stress parameters were assessed (ROS, antioxidant concentration and DNA damage), the subjects were categorized into ROS positive (ROS+) or ROS negative (ROS-). Seminal plasma from each group was pooled and subjected to proteomics analysis. In-solution digestion and protein identification with liquid chromatography tandem mass spectrometry (LC-MS/MS), followed by bioinformatics analyses was used to identify and characterize potential biomarker proteins. Results: A total of 14 proteins were identified in this analysis with 7 of these common and unique proteins were identified in both the ROS+ and ROS- groups through MASCOT and SEQUEST analyses, respectively. Prolactininduced protein was found to be more abundantly present in men with increased levels of ROS. Gene ontology annotations showed extracellular distribution of proteins with a major role in antioxidative activity and regulatory processes. Conclusions: We have identified proteins that help protect against oxidative stress and are uniquely present in the seminal plasma of the ROS- men. Men exhibiting high levels of ROS in their seminal ejaculate are likely to exhibit proteins that are either downregulated or oxidatively modified, and these could potentially contribute to male infertility. [ABSTRACT FROM AUTHOR]

Protein-protein interactions identified through high-throughput proteomics efforts continue to advance our understanding of the protein interactome. In addition to highly specific protein-protein interactions, it is becoming increasingly more common for yeast two-hybrid, pull-down assays, and other proteomics techniques to identify multiple protein ligands that bind to the same target protein. A resulting challenge is to accurately characterize the assembly of these multiprotein complexes and the competition among multiple protein ligands for a given target. The Association of Biomolecular Resource Facilities-Molecular Interactions Research Group recently conducted a benchmark study to assess participants' ability to correctly describe the interactions between two protein ligands and their target protein using primarily biosensor technologies, such as surface plasmon resonance. Participants were provided with microgram quantities of three proteins (A, B, and C) and asked to determine if a ternary A-B-C complex can form or if protein-B and protein-C bind competitively to protein-A. This article will summarize the experimental approaches taken by participants to characterize the molecular interactions, the interpretation of the data, and the results obtained using different biosensor instruments. [ABSTRACT FROM AUTHOR]