Uncovering Novel Protein Partners of Inducible Nitric Oxide Synthase in Human Testis
Karthik S. Prabhakara,
Kavya Ganapathy,
Kazi N. Islam,
Hiran M. Thyagarajan,
Kirti K. Tiwari,
Ramya L. Parimi,
Mohammad B. Rashid
Affiliations
Karthik S. Prabhakara
Department of Biology and Biotechnology, College of Science and Engineering, University of Houston-Clear Lake, 2700 Bay Area Blvd., Houston, TX 77058-1098, USA
Kavya Ganapathy
Department of Biology and Biotechnology, College of Science and Engineering, University of Houston-Clear Lake, 2700 Bay Area Blvd., Houston, TX 77058-1098, USA
Kazi N. Islam
Department of Agricultural Research and Development Program, Central State University, 1400 Brush Row Road, Wilberforce, OH 45384, USA
Hiran M. Thyagarajan
Department of Biology and Biotechnology, College of Science and Engineering, University of Houston-Clear Lake, 2700 Bay Area Blvd., Houston, TX 77058-1098, USA
Kirti K. Tiwari
Department of Biology and Biotechnology, College of Science and Engineering, University of Houston-Clear Lake, 2700 Bay Area Blvd., Houston, TX 77058-1098, USA
Ramya L. Parimi
Department of Biology and Biotechnology, College of Science and Engineering, University of Houston-Clear Lake, 2700 Bay Area Blvd., Houston, TX 77058-1098, USA
Mohammad B. Rashid
Department of Biology and Biotechnology, College of Science and Engineering, University of Houston-Clear Lake, 2700 Bay Area Blvd., Houston, TX 77058-1098, USA
Peroxidative damage to human spermatozoa has been shown to be the primary cause of male infertility. The possible role of nitric oxide (NO) in affecting sperm motility, capacitation, and acrosome reaction has been reported, too. The overproduction of NO by the enzyme inducible nitric oxide synthase (iNOS) could be responsible as it has been implicated in the pathogenesis of many diseases. There have been many studies on regulating iNOS function in various tissues, especially by protein–protein interaction; however, no study has looked for iNOS-interacting proteins in the human testis. Here, we have reported the identification of two proteins that interact with iNOS. We initially undertook a popular yeast two-hybrid assay to screen a human testis cDNA library in yeast using an iNOS-peptide fragment (amino acids 181–335) as bait. We verified our data using the mammalian chemiluminescent co-IP method; first, employing the same peptide and, then, a full-length protein co-expressed in HEK293 cells in addition to the candidate protein. In both cases, these two protein partners of iNOS were revealed: (a) sperm acrosome-associated 7 protein and (b) retinoblastoma tumor-suppressor binding protein.
