A gene coding for lysozyme from the insect Manduca sexta (Ms-lyz) was expressed in Escherichia coli. The protein was produced as an insoluble cytoplasmic inclusion body which was denatured in 8 M guanidine-HCl, renatured and purified by affinity and ion-exchange chromatography. The N-terminal sequence and the activity of the recombinant protein against Micrococcus luteus confirmed that correct expression had occurred. When Ms-lyz activity was compared to hen egg white lysozyme, the insect lysozyme was active at lower temperatures. These results demonstrate the feasibility of producing a disulfide-bonded lysozyme enzyme in bacteria and suggest that the insect Ms-lyz is an interesting system for further development of an antibacterial functional at low temperatures.
Bibliographical noteFunding Information:
This work was financed by grants 36928-B and 41184-Z from CONACyT (National Research Council of Science and Technology of México) and by grant A/3230-1 from International Foundation for Science (Sweden). A. López-Zavala and D. Puentes-Camacho received scholarships from CONACyT. We thank Drs. Maria A. Islas-Osuna and Mayra de la Torre from CIAD (Sonora, México) and Dr. Lorraine Marnell from University of New Mexico (NM, USA) for critical reading and editorial suggestions during the preparation of this manuscript.
- Affinity chromatography
- Bacterial over-expression
- Manduca sexta