Structural studies of Carboxypeptidase A thermal stabilization in the presence of spermidine

Publish Year: 1397
نوع سند: مقاله کنفرانسی
زبان: English
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شناسه ملی سند علمی:

CBC15_090

تاریخ نمایه سازی: 29 خرداد 1398

Abstract:

Carboxypeptidase A (CPA) (IUBMB: EC 3.4.17.1) is a zinc-containing proteolytic enzyme which removes the C-terminal amino acid from a peptide chain if the C-terminal carboxylate is free. Polyamines such as spermidine (NH2(CH2)3NH-(CH2)4NH2) are low-molecular-weight aliphatic polyamines that participate in cell growth, differentiation, proliferation processes and also are involved in the biology of protein aggregation and can be used to prevent thermal aggregation. In this study, the effect of spermidine on the structure and thermal stability of CPA were investigated by the method of UV-Vis, fluorescence spectroscopy and steady-state thermal denaturation at pH 7.5. It was found that by increasing of spermidine concentration, the thermal stability of CPA was increased in the range 303-353 K in the tris-HCl buffer and pH 7.5 and midpoint temperature of thermal unfolding (Tm) of CPA in the presence of spermidine, after heat treatment, was increased with increasing the concentration of this additive. The analysis of UV-Vis indicates that by increasing concentration of spermidine, absorption was decreased. In addition, Stern−Volmer quenching constants (Ksv) for the CPA−spermidine complex were obtained at two temperatures. The results of fluorescence spectroscopic measurements suggested that spermidine has a diametrical ability to quench the intrinsic fluorescence of CPA through the static quenching procedure. The thermodynamic parameters, Gibbs free-energy (ΔG°), enthalpy (ΔH°), and entropy (ΔS°) changes, shown that the binding process was spontaneous. These values revealed that the hydrogen bonding and van der Waals forces played a major role in stabilizing of this complex.

Authors

Mozhgan Mohammadi

Ph.D. Student in Biochemistry, department of biology, Faculty of Sciences, University of Shahrekord, Iran

Behzad Shareghi

Professor of Biochemistry, department of biology, Faculty of Sciences, Shahrekord, Iran

Ali Akbar Saboury

Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran