Crystal structures of pertussis toxin with NAD+ and analogs provide structural insights into the mechanism of its cytosolic ADP-ribosylation activity

• Published in: The Journal of biological chemistry Vol. 298; no. 5; p. 101892
• Main Authors: Sakari, Moona, Tran, Mai T., Rossjohn, Jamie, Pulliainen, Arto T., Beddoe, Travis, Littler, Dene R.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.05.2022; American Society for Biochemistry and Molecular Biology
• Subjects: AB5 toxin; ADP-ribosylation; Bordetella; pertussis toxin; whooping cough; ART; PT; HRP; PARP; ER; NAD; whooping cough; 3AB; Bordetella; pertussis toxin; AB5 toxin; BaAD; TCEP; PJ34; ADP-ribosylation; AB toxin
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/j.jbc.2022.101892

Bordetella pertussis is the causative agent of whooping cough, a highly contagious respiratory disease. Pertussis toxin (PT), a major virulence factor secreted by B. pertussis, is an AB5-type protein complex topologically related to cholera toxin. The PT protein complex is internalized by host cells and follows a retrograde trafficking route to the endoplasmic reticulum, where it subsequently dissociates. The released enzymatic S1 subunit is then translocated from the endoplasmic reticulum into the cytosol and subsequently ADP-ribosylates the inhibitory alpha-subunits (Gαi) of heterotrimeric G proteins, thus promoting dysregulation of G protein–coupled receptor signaling. However, the mechanistic details of the ADP-ribosylation activity of PT are not well understood. Here, we describe crystal structures of the S1 subunit in complex with nicotinamide adenine dinucleotide (NAD+), with NAD+ hydrolysis products ADP-ribose and nicotinamide, with NAD+ analog PJ34, and with a novel NAD+ analog formed upon S1 subunit crystallization with 3-amino benzamide and NAD+, which we name benzamide amino adenine dinucleotide. These crystal structures provide unprecedented insights into pre- and post-NAD+ hydrolysis steps of the ADP-ribosyltransferase activity of PT. We propose that these data may aid in rational drug design approaches and further development of PT-specific small-molecule inhibitors.