Cloning, expression and purification of collagen-like protein from Methylobacteria sp. 4-46 as alternative substance for collagen-based industries

• Main Authors: Salleh, Faqihah, Amid, Azura, Nordin, Noor Faizul Hadry
• Format: Conference Object
• Language: English
• Published: International Institute for Halal Research and Training (INHART), International Islamic University Malaysia (IIUM) 2021
• Subjects: Q Science (General); Haram; Avian flu
• Online Access: http://irep.iium.edu.my/92122/; http://irep.iium.edu.my/92122/2/92122%20Cloning,%20expression%20and%20purification%20of%20collagen-like%20protein.pdf

Nowadays, collagen is widely introduced into oral and topical relief cosmeceutical products and is applied in the pharmaceutical industry in the form of hydrogels and collagenated scaffolds. However, due to the source of collagen in the market, it is a big concern for the Muslim community in terms of Halalan and Toyyiban since it is mainly extracted from mammals, especially porcine (Haram) and it raises health issues of animal-to-human disease transmission such as mad cow disease, swine, and avian flu. Collagen from bacteria would be the best alternative for halal and non-animal collagen. In this study, the bacterial collagen that produces collagen-like protein (CLP) of Methylobacteria sp. 4-46 was synthetically synthesized by Genscript. The CLPM4-46 in pUC57 was transformed into Escherichia coli DH5α and digested with BamHI/NdeI. The digested product (CLPM4-46, 1.04kbp) was successfully ligated with pCold II plasmid (4.39kbp) and transformed into E. coli BL21 (DE3). The ligated CLP gene was expressed using M9 minimal media. The protein expressed by the recombinant E. coli was measured with the Bradford Assay and SDS-PAGE where the size of the protein was 34.9k Da, and the CLP recombinant protein was purified using Fast Protein Liquid Chromatography and the collagen was measured using Enzyfluo™ Collagen Assay