PJB-2018-299
Structural characterization of 12S seed Cruciferin from Eruca sativa in solution applying small-angle X-ray scattering
Binish Khaliq, Sven Falke, Sadaf Iqbal, Friedrich Buck, Seema Mahmood, Zahid Ali, Maqsood Ahmad, Abdul Wahid Christian Betzel and Ahmed Akrem
DOI:10.30848/PJB2019-4(29)
Abstract
Cruciferin (12S globulin) from seeds of Eruca sativa was isolated, purified and thoroughly characterized. The protein indicated 54 % sequence identity with the cruciferins of Brassica napus (CRU1) and Raphanus sativus (PGCRURSE5) when the obtained amino acid sequence from LC-MS/MS mass spectrometric data was submitted to the UniProtKB. SDS-PAGE exhibited an approx. 50 kDa monomeric cruciferin, which was separated into α-polypeptide with a major band at approx. 30 kDa and a β-polypeptide of approx. 20 kDa under reduced conditions. The secondary structure content of E. sativa Cruciferin (EsC) was analyzed by Circular Dichroism spectroscopy indicating 7% α-helix, 48% β-sheet, 7% β-turn and 38% disordered conformation. The monodispersity and stability of EsC was verified via Dynamic Light Scattering (DLS) and a hydrodynamic radius of EsC was calculated to be 5.5 ± 0.3 nm indicating a trimer of the protein in solution. A gyration radius (Rg) of 4.3± 0.30 nm and the globular molecular shape was disclosed by Small-angle X-ray scattering (SAXS) for EsC. An incredibly analogous globular shape was obtained when the ab-initio dummy model of EsC inferring P3 symmetry was carefully compared with the PDB-ID 3KGL; 11S globulin of Brassica napus. Moreover, the scattering patterns of both proteins showed a minimized χ2-value of 2.0 which further confirms the structural similarities. Protruding loops of the EsC model were considered as hyper variable region-I (HVR-I) of Arabidopsis thaliana Cruciferin C (AtCruC) and variable region II of 3KGL molecular structure and were nominated as the major flexible regions.
To Cite this article:
Khaliq, B., S. Falke, S. Iqbal, F. Buck, S. Mahmood, Z. Ali, M. Ahmad and A. Wahid. 2019. Christian Betzel and Ahmed Akrem - Structural characterization of 12S seed Cruciferin from Eruca sativa in solution applying small-angle X-ray scattering. Pak. J. Bot., 51(4): DOI: http://dx.doi.org/10.30848/PJB2019-4(29)
Download