Purification and crystallization of I-SSC1
Research Mentor(s)
Spiegel, P. Clint
Description
Homing endonucleases are a class of enzymes capable of hydrolyzing DNA within the nucleus of the cell that synthesized them. As the DNA is repaired the sequence encoding the endonuclease is often copied to the sight of cleavage. These target DNA sequences are commonly 12 to 40 base pairs long and differentiate the endonucleases into families. The 300 amino acid protein, I-SSC1 is an intron-encoded protein within the LAGLIDADG family, named from their target sequences. To date there are no proposed structures for this protein. We have been working on the purification and crystallization of a His-tagged version of I-SSC1. Expression and purification methods have been successfully determined using E. coli as a means of expression, and immobilized metal ion affinity chromatography (IMAC) for the purification. Crystallization conditions are still being optimized, but initial crystals have been grown.
Document Type
Event
Start Date
May 2018
End Date
May 2018
Department
Chemistry
Genre/Form
student projects, posters
Subjects – Topical (LCSH)
Enzymes--Biotechnology; Protein engineering; Human genetics; Genetic engineering
Type
Image
Rights
Copying of this document in whole or in part is allowable only for scholarly purposes. It is understood, however, that any copying or publication of this document for commercial purposes, or for financial gain, shall not be allowed without the author’s written permission.
Language
English
Format
application/pdf
Purification and crystallization of I-SSC1
Homing endonucleases are a class of enzymes capable of hydrolyzing DNA within the nucleus of the cell that synthesized them. As the DNA is repaired the sequence encoding the endonuclease is often copied to the sight of cleavage. These target DNA sequences are commonly 12 to 40 base pairs long and differentiate the endonucleases into families. The 300 amino acid protein, I-SSC1 is an intron-encoded protein within the LAGLIDADG family, named from their target sequences. To date there are no proposed structures for this protein. We have been working on the purification and crystallization of a His-tagged version of I-SSC1. Expression and purification methods have been successfully determined using E. coli as a means of expression, and immobilized metal ion affinity chromatography (IMAC) for the purification. Crystallization conditions are still being optimized, but initial crystals have been grown.