Clostridial Sources

Whereas the use of synthetic substrates in lieu of native collagen is appropriate in studies with collagenase from clostridial sources, their use in studies of collagenases derived from vertebrate sources cannot be endorsed at this time. The actual amino acid sequence and or other structural requirements that may be recognized by vertebrate collagenases are still unknown. Until these requisites have been determined, we will not know whether splitting of a synthetic substrate represents nonsi)Ccific peptidase activity or specific collagenolytic effect. 

Fig. 11. Mossbauer spectra at low temperature and small applied magnetic field for reduced plant-type ferredoxins. Abbreviations AZI = Azotobacter Fe-S Protein I, 4.2 °K, 1.15 kG AZIl — Azotobacter Fe-S Protein II, 4.2 °K, 300 G Put. = Putid-aredoxin, 4.6 °K, 580 G Clos. = Clostridial Paramagnetic Protein, 4.7 K, 3.4 kG Ad. = Pig Adrenodoxin, lyophilized, 5.3 C>K, 580 G PPNR = Spinach Ferredoxin, lyophilized, 4.3 °K, 580 G Parsley = Parsley Ferredoxin, 5.1 °K, 580 G. Applied magnetic field is parallel to gamma-ray direction. Velocities are relative to platinum source matrix...

Niemann H (1991) Molecular biology of clostridial neurotoxins. In Alouf J, Freer J (eds) A source-book of bacterial protein toxins. Academic Press, London, pp 303 18 Nishiki T, Tokuyama Y, Kamata Y, Nemoto Y, Yoshida A et al. (1996) The high-affinity binding of Clostridium botulinum type b neurotoxin to synaptotagmin ii associated with gangliosides gtlb/gdla. FEBS Lett 378 253-7... 

Bayer et al. (7) succeeded in reconstituting clostridial ferredoxin from a, a -dipyridyl-treated apoferredoxin in the presence of ferrous ion without the presence of a labile sulfur source. Based on this observation, they proposed that labile sulfur originates from cysteinyl residues in the presence of iron. Contrary to their finding, Rabinowitz and his coworkers (21, 33) demonstrated that the reconstitution of clostridial ferredoxin from mersaryl-, a, a -dipyridyl-, and trichloroacetic acid-treated apo-ferredoxins absolutely requires the addition of hydrogen sulfide. Consequently, they concluded that cysteine is not the source of labile sulfur . 

Although the mechanism of action of 2-mercaptoethanol may not be simple, its probable action is the reduction of disulfide bonds to sulfhy-dryl groups in a, a -dipyridyl- or heat-treated apoadrenodoxin, or regeneration of sulfhydryl groups from mercaptides in PCMB-treated apoadrenodoxin. At the present time, all three laboratories, that is Bayer s Rabinowitz s, and ours, agree on the necessity of the addition of Na2S as a labile sulfur source for better or perfect reconstitution of clostridial, spinach, and adrenal non-heme iron proteins from various types of apoproteins. 

FHatheway CL (1990) Bacterial Sources of Clostridial Neurotoxins. In Botulinum Neurotoxin and Tetanus Toxin. (Simpson, LL, ed) pp 3-24, San Diego Academic Press... 

---