Pterin-containing molybdenum enzymes catalyze a wide variety of reactions and those that have been overexpressed focused on the catalytic subunit. In select nitrate reductases, the chaparones and maturation proteins are absolutely vital in rendering the catalytic subunit functional. In bacterial periplasmic nitrate reductase, (e.g., from Campylobacter jejuni) the catalytic subunit, NapA, matures with the help of other smaller subunits. While NapA is known to play a role in the infectious cycle of C. jejuni, precious little is known about these important maturation proteins.  Here we report the cloning of two of those suspected proteins, NapD and NapL, into separate pET based expression vectors and modified with a 6x-poly His tag for use in purification.  Furthermore, we discuss the overexpression via IPTG induction in the commonly used E. coli expression strain, BL21(DE3).  The expressed proteins have been characterized by SDS-PAGE and LC/MS-MS.

Heterologous cloning and overexpression of periplasmic nitrate reductase maturation proteins, NapL and NapD

247th National Meeting (2014)

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