Biophysical characterization of the structural stability of Helix lucorum hemocyanin

• Main Authors: Krassimira Idakieva, Svetla Todinova, Aleksandar Dolashki, Lyudmila Velkova, Yuliana Raynova, Pavlina Dolashka
• Format: Article
• Language: English
• Published: Taylor & Francis Group 2021-01-01
• Series: Biotechnology & Biotechnological Equipment
• Subjects: hemocyanin; mollusk; helix lucorum; thermal stability
• Online Access: http://dx.doi.org/10.1080/13102818.2020.1837010
• ISSN: 1310-2818; 1314-3530

The structural stability of the hеmocyanin purified from the hеmolymph of gаrden snаils Helix lucorum (HlH) was investigated by means of far-UV circular dichroism (CD), differential scanning calorimetry (DSC) and transmission electron microscopy (TEM). For the first time, TEM analyses showed the presence of tubular polymers in hemocyanins after three-day dialysis against a stabilizing buffer containing high concentrations of Ca2+ and Mg2+ ions (100 mmol L−1). The conformational stability study of native HlH by means of CD in a wide pH range (2.5-11.5) defined the pH stability region of HlH at pH 6.5 − 8.0. DSC analyses demonstrated the thermal stability of this hemocyanin. One transition, with an apparent transition tеmperature (Tm) at 82.3 °C, was detected in the heаt capacity curve of HlH in 50 mmol L−1 Tris-HCl buffer, pH 7.2, at a heating rate of 1.0 °C min−1. The calorimetrically observed thermal transition correlates well with the unfolding transition monitored by CD measurements. The two-state kinetic model was used to analyse the process of irreversible thermal denaturation of HlH; Еa of 451 ± 4 kJ mol−1 was calculated. The obtained results on the conformational stability of HlH will facilitate the further investigation of the properties and potential biomedical applications of this respiratory protein.