Human ribonuclease 9, a member of ribonuclease A superfamily, specifically expressed in epididymis, is a novel sperm-binding protein

Abstract

To explore the functions of human ribonuclease 9 (RNase 9), we constructed a mammalian fusion expression vector pcDNA-hRNase9, prepared recombinant human RNase 9-His fusion protein from HEK293T cells and determined its N-terminal amino acid sequences. According to the determined mature protein, recombinant human RNase 9 was prepared in E. coli. Ribonucleolytic activity and antibacterial activity of recombinant human RNase 9 were detected, and the distribution of human RNase 9 on tissues and ejaculated spermatozoa and in vitro capacitated spermatozoa were analyzed via indirect immunofluorescence assay. The results showed that recombinant human RNase 9 did not exhibit detectable ribonucleolytic activity against yeast tRNA, but exhibited antibacterial activity, in a concentration/time dependent manner, against E. coli. Immunofluorescent analyses showed that the predicted human RNase 9 was present throughout the epididymis, but not present in other tissues examined, and human RNase 9 was also present on the entire head and neck regions of human ejaculated spermatozoa and in vitro capacitated spermatozoa. These results suggest that human RNase 9 may play roles in host defense of male reproductive tract.

Characterization of the porcine seminal plasma proteome comparing ejaculate portions

“…Other of these SP-proteins more expressed in P3, as Alpha-enolase or Alkaline phosphatase (ALP), have been related to sperm motility [52][53]. Noticeable, deoxyribonuclease -2-alpha, an acid endonuclease secreted by male accessory glands, is involved in the degradation of exogenous DNA [54] and it also provide a bactericide activity protecting sperm in the transit along female genital tract [55]. Nucleobinding-1 belongs to a family of proteins with calcium and DNA binding properties [56], but its reproductive function is still unknown.…”

In SilicoPrediction andIn VitroCharacterization of Multifunctional Human RNase3

“…To date, 13 members of the RNaseA superfamily have been identified in humans; however, the functions of newly identified human hRNases9–13 remain unclear [53]. Blast analysis of HBP RNase3(32–41) motif among hRNase3 and other hRNaseA members was shown in Table 5, in which only hRNase2 and hRNase8 showed, respectively, 80% and 50% sequence identity, while the others showed lower than 50% identity with the HBP RNase3(32–41) of hRNase3.…”

Prediction of Regulatory Molecules for Enhancement of EGFR Overexpression Triggered by Signal Peptide of Ribonuclease 3

“…Up to now, nine human proteins with significant sequence similarities to bovine pancreatic RNaseA, the first isolated RNase, are grouped into human RNaseA superfamily and are named as RNase1 to RNase9 [2][3][4]. Within this protein superfamily, two categories can be classified; one possesses high ribonucleolytic activity and the other is cytotoxic toward bacteria, parasites, or mammalian cells [3,[5][6][7]. Human RNase3, also named as eosinophil cationic protein (ECP), is one of the cytotoxic RNases whose common features include high isoelectric point (pI) and interaction with negatively charged cellular components such as heparan sulfate and lipid on cell membrane [8,9].…”

“…Up to now, nine human proteins with significant sequence similarities to bovine pancreatic RNaseA, the first isolated RNase, are grouped into human RNaseA superfamily and are named as RNase1 to RNase9 [2][3][4]. Within this protein superfamily, two categories can be classified; one possesses high ribonucleolytic activity and the other is cytotoxic toward bacteria, parasites, or mammalian cells [3,[5][6][7].…”