Skip to: main navigation | content

SLAC National Accelerator Laboratory

Botulinum Neurotoxin is Bio-shielded by NTNHA in a Handshake Complex
SSRL Science Summary - October 2012

A single reconstructed slice and a volume rendering of the tomography sequence.

Botulinum neurotoxins (BoNTs) invade motor neurons at their junctions with muscular tissue, where the toxins disable the release of the neurotransmitter acetylcholine and subsequently paralyze the affected muscles. Accidental BoNT poisoning primarily occurs through ingestion of food products contaminated by Clostridium botulinum, the bacterium that produces BoNTs. However, BoNTs by themselves are fragile and sensitive to low pH environments and digestive proteases. So how do they survive the harsh environment of the host's gastrointestinal tract?

Researchers at Sanford-Burnham Research Institute and the Medical School of Hannover in Germany recently discovered that the neurotoxin has a bodyguard, a protein termed non-toxic non-hemagglutinin (NTNHA) protein that is produced by the C. botulinum bacterium simultaneously with BoNT. This study, published in Science, revealed the first crystal structure of a ~300kDa protein complex composed of serotype A BoNT (BoNT/A) and NTNHA, which together form the minimally functional progenitor toxin complex (M-PTC) (Figure 1). A genetically modified, catalytically inactive BoNT/A that carries three mutations in its catalytic site (termed BoNT/Ai) was used for the structural studies. The M-PTC crystal structure was determined at 2.7 Å resolution by single isomorphous replacement with anomalous scattering (SIRAS) method using x-ray diffraction data collected at SSRL's Beam Line 9-2 and NE-CAT at the Advanced Photon Source. Surprisingly, the non-toxic NTNHA exhibited a three-domain architecture similar to the neurotoxin, although the two proteins share less than 20% sequence identity (Figure 1). The two proteins hug each other, interlocking with what looks like a handshake and providing unusually large and multivalent binding interfaces that shield BoNT from the hostile gastrointestinal environment. Biochemical and functional studies also revealed a novel pH-sensing mechanism which enables the M-PTC to remain tightly bound at acidic pH in the gut, dissociating at neutral pH after absorption from the gut into the general circulation.

Several essential residues involved in pH-dependent assembly of BoNT/A and NTNHA, which are weak spots of the toxin that could be targeted with new therapeutics, were identified by structure-based mutagenesis. This discovery suggests an innovative approach for oral inhibitor design that aims to mimic pH change by prematurely breaking up BoNT's protective embrace and leaving the unprotected toxin to be targeted by the stomach's digestive enzymes and acid. This type of therapy-either alone or in combination with other therapies currently in development-could be given preventively at a time when BoNT contamination becomes a public health concern. Furthermore, a better understanding of the toxin protection mechanism could enable the design of novel proteinaceous vehicles to allow oral administration of drugs that currently must be injected.


Primary Citation

Shenyan Gu, Sophie Rumpel, Jie Zhou, Jasmin Strotmeier, Hans Bigalke, Kay Perry, Greice Krautz-Peterson, Charles B. Shoemaker, Andreas Rummel, Rongsheng Jin. (2012) "Botulinum neurotoxin is shielded by NTNHA in an interlock complex." Science, 335 (6071): 977-981. [DOI: 10.1126/science.1214270]

Related Links


Rongsheng Jin, The Sanford-Burnham Research Institute

2575 Sand Hill Road, MS: 99, Menlo Park, California, 94025, USA Tel: 650-926-4000 | Fax: 650-926-4100