Structural heterogeneity, post-translational modifications, and biological activities of SV-IV, a major protein secreted from the rat seminal vesicle epithelium

Abstract

The primary structure of purified SV-IV, a major secretory protein synthesized by the rat seminal vesicle (SV) epithelium, was analysed by electrospray mass spectrometry (ES-MS). The protein was found to be highly heterogeneous. The various components were separated and identified by reversed phase high-performance liquid chromatography (HPLC) on line with ES-MS. Structural characterization of the SV-IV cyanogen bromide digests revealed the occurrence of a Val/Met substitution in about 50% of the purified protein molecules. We suggest that this mutation is the expression of a genetic polymorphism. Other minor components, corresponding to structural changes (fragmentation, deletion, and phosphorylation) of SV-IV and probably due to post-translational modifications of the native protein, were also detected. In particular, by using protein tyrosine phosphatase hydrolysis combined with ES-MS, we demonstrated that, in the phosphorylated species of SV-IV, a single phosphate group was covalently bound to the Tyr-36 residue. The significance of these findings in relation to the regulation of important biological processes, such as immune response, blood coagulation, inflammatory reaction, and mammalian reproduction, are discussed.

The N-terminal 1-16 peptide derived in vivo from protein seminal vesicle protein IV modulates α-thrombin activity: potential clinical implications

“…This finding suggests a physiological role of P1-16 in the homeostatic control. It was reported that the semen contains a thrombin-like enzyme, prothrombin fragments 1 and 2 (F1 + 2), D-dimer (DD) and thrombin-AT (TAT) complexes as well as the seminal vesicle proteins (Ferranti, 1997). These proteins may have roles in seminal clotting and in liquefaction through "fibrinolytic" activity, which may ultimately affect fertility.…”

“…We have previously shown with hydrolytic experiments that the modulating effect on the coagulation of the protein is located in its 1-70 N-terminal fragment (Di Micco, 1994. Mass spectrometry analysis of purified SV-IV demonstrated that it is highly heterogeneous (Ferranti, 1997). Truncated forms of SV-IV are also present and represent about 14% of the total molecules, in detail the 1-16, 1-17, and 1-18 peptides derived from the N-terminus of the protein.…”

“…Truncated forms of SV-IV are also present and represent about 14% of the total molecules, in detail the 1-16, 1-17, and 1-18 peptides derived from the N-terminus of the protein. The processing mechanism leading to their formation could be related to the synergistic effects of chymotrypsinlike and carboxypeptidase enzymes (Ferranti, 1997).…”

Conformational Analysis of Rat Seminal Vesicle Secretory Protein 4, an Intrinsically Disordered Protein Having Interesting Pharmacological Properties

“…Many studies on the RSV4 (Ferranti et al, 1997;Metafora et al, 2001;Vilasi and Ragone, 2008;Abrescia et al, 1986;Porta et al, 1990;Di Micco et al, 2000;Stiuso et al, 1999;Porta et al, 1994;Metaofra et al, 1987;Suskiewicz et al, 2011) were the result of the collaboration for many years of several groups operating in Naples (Italy). This protein has always shown resistance to reveal details of its three-dimensional structure, despite being attempted many times its crystallization but with no results.…”

Structural properties of the protein SV‐IV

“…We have demonstrated recently by electrospray MS that 10% of the native SV‐IV molecules are phosphorylated in vitro by protein kinase C and that this modification involves only Ser58 [23]. Furthermore, we have unambiguously demonstrated that a Tyr36‐linked phosphate group is present in 14% of all native SV‐IV molecules [24].…”