Secondary structure of protamine in sperm nuclei: an infrared spectroscopy study

Abstract

BackgroundProtamines are small basic proteins that condense the DNA in mature spermatozoa. Typical protamines are of simple composition and very arginine-rich, usually in the range of 60-80%. Arginine residues are distributed in a number of stretches separated by neutral amino acids. We have used Fourier transform infrared spectroscopy (FTIR) to gain access for the first time to the secondary structure of protamines in sperm nuclei. This technique is particularly well suited to the study of DNA-bound protamine in whole nuclei since it is not affected by turbidity.ResultsWe show that DNA -bound salmon (salmine) and squid protamines contain α-helix, β-turns and a proportion of other structures not stabilized by intramolecular hydrogen bonding. No β-sheet was observed. In salmine, the α-helix amounted to ~20%, while in squid protamine it reached ~40%. In contrast, the structure not stabilized by intermolecular hydrogen bonding was more abundant in salmine (~40%) than in squid protamine (~20%). Both protamines contained ~40% β-turns. The different helical potential of salmine and squid protamine was confirmed by structure predictions and CD in the presence of trifluoroethanol.ConclusionDNA-bound protamine in sperm nuclei contains large amounts of defined secondary structure stabilized by intramolecular hydrogen bonding. Both salmine and squid protamine contain similar amounts of β-turns, but differ in the proportions of α-helix and non-hydrogen bonded conformations. In spite of the large differences in the proportions of secondary structure motifs between salmon and squid protamines, they appear to be equally efficient in promoting tight hexagonal packing of the DNA molecules in sperm nuclei.

Fourier transform infrared spectroscopic analysis of sperm chromatin structure andDNAstability

“…Microspectroscopic approaches have been used to assign spectral properties that are typical for normal or abnormal and damaged spermatozoa (Kubasek et al ., ; Huser et al ., ; Meister et al ., ; Mallidis et al ., ; Sánchez et al ., ). Spectral fingerprints of cells are complex, because vibration bands of characteristic biomolecular groups often overlap and are not necessarily the same as in model systems (Hud et al ., ; Roque et al ., ; Mello & Vidal, ). Studies on isolated components (e.g., DNA, protamines), evaluation of medium contributions (e.g., using deuterated water, or confocal Raman imaging), as well as analysis of samples with defined damage (e.g., oxidative damage) have proven to be useful to assign spectral features (Mello & Vidal, ; Sánchez et al ., ; D'Amico et al ., ).…”

Paternal contribution to development: Sperm genetic damage and repair in fish

“…Protamines are a group of highly diverse arginine (and sometimes cysteine)-rich proteins (> 30% arginine) with a small size that can range from 30-100 amino acids (Kasinsky et al 2011). Structurally these are intrinsically disordered proteins (IDPs) (Dunker et al 2001) that do not exhibit any structural features in solution, but can adopt highly organized structures upon interaction with DNA (Roque et al 2011). These proteins displace and completely replace the somatic histone counterpart over the course of spermiogenesis.…”

MicroRNA-122 Influences the Development of Sperm Abnormalities from Human Induced Pluripotent Stem Cells by Regulating TNP2 Expression

“…They are characterized by a number of arginine residue stretches separated by neutral amino acids. Fiber-diffraction diagrams from reconstituted nucleoprotamine and whole sperm cells indicated that the DNA molecules were tightly packed in a hexagonal unit cell, and that DNA was in a B-like structure with 10 base pairs per helical turn [18]. PRM1 and PRM2, the two PRMs found in mammals, are the most widely studied.…”

“…PRM2 (but not PRM1) is synthesized as a precursor that undergoes proteolytic processing after binding to DNA. It also binds to a zinc atom, albeit its function is not yet known [17,18].…”

“…Furthermore, other studies have demonstrated the importance of PRMs in the morphogenesis and function of mature spermatozoa [17,18]. PRMs are a diverse family of small arginine-rich proteins that have been found to be synthesized in the late-stage spermatids of many animals and plants.…”