Clostridium Stability

Within the cellulosome complex, type I dockerin domain is responsible for incorporating its associated glycosyl hydrolase in the bacterial cellulosome via interaction with a reception domain, the cohesin domain. The three-dimensional solution structure of the 69-residue dockerin domain from the thermophilic Clostridium thermocellum (Topt = 55-65 °C) was solved by NMR and was found to consist of two Ca " -binding loop-helix motifs connected by a linker. Each Ca " -binding subdomain is stabilized by a cluster of buried hydrophobic sidechains. Recently, the NMR sequence-specific resonance assignment of type II cohesin module from C. thermocellum has been published. ... 

Halouzka, J., Hubalek, Z. (1992). Effect of pH on the stability of type-C toxin of Clostridium botulinum. Folia Microbiol. (Praha) 37 157-8. 

Sugii, S., Ohishi, L, Sakaguchi, G. (1977a). Correlation between oral toxicity and in vitro stability of Clostridium botulinum type A and B toxins of different molecular sizes. Infect. Immun. 16 910-14. 

Detoxification. The process by which bacterial toxins are converted to harmless toxoids. Formaldehyde is used to detoxify the toxins of Corynebacterium diphtheriae, Clostridium botu-linum and Cl. tetani. The detoxification may be performed either on the whole culture in the fermenter or on the purified toxin after fractionation. Traditionally the former approach has been adopted, as it is much safer for the operator. However, the latter gives a purer product. The pertussis toxin used in acellular vaccines may be detoxified with formaldehyde, glutaraldehyde, or both, hydrogen peroxide or tetranitromethane. In the case of genetically detoxified pertussis toxin, a treatment with a low concentration of formaldehyde is still performed to stabilize the protein. 

LAM from Clostridium subterminale in complex with the substrate crystallizes as a homotetramer with domain-swapped dimers stabilized by structural zinc ions. Each monomer contains one active site, which is a channel defined by a P / a) -crescent and binds a molecule of PLP, a molecule of SAM, and a [4Fe-4S] cluster. In the active site the external aldimine of PLP with lysine, SAM, and the Fe-S cluster lie spatially close (Figure 29), supporting the idea that the formation of the 5 -deoxyadenosyl radical and the abstraction of the... 

Stabilities of GDH s from microorganisms vary greatly. Whereas the NADP-GDH from Neurospora retains activity for several days at 50° and pH 7.2 (152), the NAD-GDH inactivates rapidly at raised temperatures and low ionic strengths, but is more stable in the presence of NAD (1 mAf) and the competitive inhibitor isophthalate (64). NAD-GDH s from lower fungi are also reported to be unstable (71), but those isolated from Clostridium SBj (35) and Peptococcus aerogenes (31) are stable at 50°, and so are the NADP-dependent enzymes of Salmonella... 

Table IV. Enzymatic Activity and Stability of Modified Ferredoxins Prepared from Clostridium aciduurici Apoferredoxin Derivatives...

Grafting of CBM has also been shown to alter the activity and themaal stability of the catalytic domain in some cases. Fusion of CBM4 to the feruloyl acetyl esterase domain (FAE) of Clostridium stercorarium XynZ was shown to result in a drastic decrease in die diermal stability of the FAE domain (7). On the other hand, Karita et al. (8) reported improvement in the thermal stability of R. albus endoglucanase upon fusion widi a fiunily 6 CBM. 

Mori and Inaba (1990) applied a PV technique to attain both high productivity and efficient recovery of EtOH from a fermentation broth. The manbrane bioreactor consisted of a jar fermenter and a PV systan for the direct production of EtOH from uncooked starch with a thermophilic anaerobic bacterium, Clostridium thermohydrosulfuricum. From the four types of EtOH-selective monbranes tested, microporous PTFE membrane, the pores of which were impregnated with silicone rubber, was chosen for its large flux, high EtOH selectivity, and high stability. During... 

Yu MR, Du YM, Jiang WY, Chang WL, Yang ST, Tang IC. (2012). Effectsof differentreplicons in conjugative plasmids on transformation efficiency, plasmid stability, gene expression and n-butanol biosynthesis in Clostridium tyrobutyricum. Appl Microbiol Biotechnol, 93, 881-889. 

Andrade JC, Vasconcelos I. (2003). Continuous cultures of Clostridium acetobutylicum culture stability and low-grade glycerol utilisation. Biotechnol Lett, 25,121-125. 

Figure 1. Effect of temperature on stability and activity of Clostridium thermohydrosulfUriciun strain 39E amylopuUulanase. (a) TherW stability. The enzyme was placed in acetate buffer (SO mM, pH 6.0) with 5 mM CaC and preincubated at various temperatures for 30 min, and then residual amylopuUulanase activities were assayed, (b) Effect of heat on activi. The enzyme activity was assayed at various temperatures by the standard assay method (30 min incubation). (Reproduced with permission from Ref. 55. Copyright 1988, The Biochemical Society and Pordand Press, London.)...

Recently, a fusion protein between a cellulose binding domain (CBOeios)> isolated from Clostridium cellulovorans, and OPH was shown to be capable of immobilization onto various cellulose materials (24). The use of cellulose as an immobilization matrix is advantageous due to its low cost, wide spread availability and non-toxic nature. The kinetic parameters of OPH fused to die CBD domain were essentially identical to die soluble protein with a 3.8 fold increase in from 0.058 to 0.220 and a 10.4% decrease in Kd from 3170 to 2840. Additionally, the immobilized fusion protein offered superior stability over that of soluble OPH, retaining over 85% activity over a period of 45 days (24). 

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