Date on Honors Thesis

Spring 5-2020


**Jesse D. Jones College of Science, Engineering and Technology**





Examining Committee Member

Chris Trzepacz, PhD, Committee Member

Examining Committee Member

Ricky Cox, PhD, Committee Member

Examining Committee Member

Christopher Lennon, PhD, Advisor


Inteins are emerging as post-translational regulatory elements, undergoing conditional protein splicing in response to a variety of environmental cues. Inteins are seamlessly removed by self-splicing from exteins, or flanking portions of the host protein, which they interrupt. DnaB of Mycobacterium smegmatis, a helicase essential for replication, harbors two inteins known as DnaBi1 and DnaBi2, each with discrete structural characteristics and insertion positions. DnaBi1 was used here to design a reporter system which links splicing with resistance to the antibiotic kanamycin. We built a construct that strictly requires catalytically active DnaBi1 for survival in the presence of kanamycin and used this construct to probe for factors that influence splicing in vivo of M. smegmatis, the native context of the intein. We show that zinc, biologically relevant during mycobacterial pathogenesis, is a potent inhibitor of DnaBi1 in vivo, and using a separate reporter system, that zinc inhibits DnaBi1 in vitro. We present the crystal structure of DnaBi1 bound to zinc, and finally propose that DnaBi1 splicing inhibition by zinc may be important during mycobacterial infection.