Structure of Clostridial Neurotoxins

The mechanism of neurotoxin cell intoxication consists of four distinct steps (a) binding to the neuronal membrane, (b) internalization into an acidic compartment, (c) membrane translocation and (d) enzymatic target modification (Montecucco etal., 1994). The L chain is responsible for the intracellular catalytic activity of CNTs. The amino-terminal 50 kDa domain of the H chain (Hn) is implicated in membrane translocation, while the carboxy-terminal part (He) is mainly responsible for the neurospecific binding. 

The amino acid sequence of all eight CNTs has been derived from their corresponding genes (Minton, 1995). The L chains and H chains are composed on average of 439 and 843 residues, respectively. Both chains contain homologous domains separated by regions of very little similarity (Fig. 2). The most conserved portions of the L chains are the amino-terminal and central regions (residues 216-244, 

BoNT/C LMHEtNHAMHNLYG BoNT/D LMHELTHSLHQLYG BoNT/E LMHELIHSLHGLYG 

In order to determine the number of histidine residues involved in zinc coordination, the L chains of TeTx and BoNT/A, B and E were modified with diethyl pyrocarbonate (DEPC), a reagent that specifically modifies histidine residues. In each case, two additional histidines were modified in the apo-toxin that were not affected in the holo-neurotoxin (Schiavo etal., 1992 b, c). These results indicate that the zinc atom of CNTs is coordinated via two histidines and a Glu-bound water molecule, as in thermolysin. Mutations at the two histidines of the motif inactivate TeTx and suppress its ability to bind radio-labeled Zn (Yamasaki etal., 1994 b). In addition, mutations of the conserved Glu-271 and Glu-272 of TeTx, predicted to be in an a-helical segment (Lebeda and Olson, 1994), result in decreased zinc binding and loss of activity. Based on these experimental results, it has 

The H chains are less conserved than the L chains and the carboxyl-terminal part of the H chain (He) is the most variable region of the toxin (Fig. 2) (Minton, 1995). This is consistent with the notions that the He domain is involved in binding to the nerve terminals and that different neurotoxins bind to different cognate receptors. On this basis it may be suggested that the receptor binding regions of TeTx and BoNTs are mainly located within the 180 carboxy-terminal residues of the H chain. 

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