Biochemical and conformational characterisation of HSP‐3, a stallion seminal plasma protein of the cysteine‐rich secretory protein (CRISP) family

FEBS Letters

Gasset, Marı́a²  Töpfer-Petersen, Edda³  Varea, Julio²  Urbanke, Claus⁴  Calvete, Juan J²  Magdaleno, Leticia²  Raida, Manfred¹  Schambony, Alexandra M³ 

[1] Niedersächsisches Institut für Peptid-Forschung (IPF) GmbH, Hannover, Germany;Instituto de Quı́mica-Fı́sica `Rocasolano', C.S.I.C., Madrid, Spain;Institut für Reproduktionsmedizin, Tierärztliche Hochschule, Bünteweg 15, D-30559 Hannover-Kirchrode, Germany;Medizinische Hochschule, Meßgeräteabteilung, Hannover, Germany

关键词: Horse seminal plasma protein;    HSP-3;    Cysteine-rich secretory protein;    Mass spectrometry;    Conformational analysis;   

DOI  :  10.1016/S0014-5793(97)01514-7

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

HSP-3 is a member of the cysteine-rich secretory protein (CRISP) family from stallion seminal plasma. We report a large-scale purification protocol for native HSP-3. This protein is a non-glycosylated polypeptide chain with a pI of 8–9 and an isotope-averaged molecular mass of 24 987±3 Da. The molecular mass of HSP-3, determined by equilibrium sedimentation, is 26 kDa, showing that the protein exists in solution as a monomer. The concentration of HSP-3 in the seminal plasma of different stallions ranged from 0.3 to 1.3 mg/ml. On average, 0.9–9 million HSP-3 molecules/cell coat the postacrosomal and mid-piece regions of an ejaculated, washed stallion spermatozoon, suggesting a role in sperm physiology. Conformational characterisation of purified HSP-3 was assessed by combination of circular dichroism and Fourier-transform infrared spectroscopies and differential scanning microcalorimetry. Based on secondary structure assignment, HSP-3 may belong to the α+β class of proteins. Thermal denaturation of HSP-3 is irreversible and follows a non-two state transition characterised by a T _m of 64°C, an enthalpy change of 75 kcal/mol, and a van 't Hoff enthalpy of 184 kcal/mol. Analysis of the spectroscopic and calorimetric data indicates the occurrence of aggregation of denatured HSP-3 molecules and suggests the monomer as the cooperative unfolding unit.

【 授权许可】

Unknown