Sequencing and Expression of a β-Mannanase Gene from the Extreme Thermophile Dictyoglomus thermophilum Rt46B.1, and Characteristics of the Recombinant Enzyme

A β-mannanase gene (manA) was isolated from the extremely thermophilic bacterium Dictyoglomus thermophilum Rt46B.1. ManA is a single-domain enzyme related to one group of β-mannanases (glycosyl hydrolase family 26). The manA gene was expressed in the heat-inducible vector pJLA602 and the expression product, ManA, purified to homogeneity. The recombinant ManA is a monomeric enzyme with a molecular mass of 40 kDa and an optimal temperature and pH for activity of 80°C and 5.0. In the absence of substrate, the enzyme showed no loss of activity at 80°C over 16 h, while at 90°C the enzyme had a half-life of 5.4 min. Hydrolysis of the galactomannan locust bean gum (LBG) by purified ManA released mainly mannose, mannobiose, and mannotriose, confirming that ManA is an endo-acting β-mannanase. Sequence comparisons with related β-mannanases has allowed the design of consensus PCR primers for the identification and isolation of related genes. Received: 7 June 1999 / Accepted: 6 July 1999     

Feather damage due to mycotic infections in wild turkeys

Wild turkeys (Meleagris gallopavo) from Pearl River Wildlife Management Area, St. Tammany Parish and from adjacent St. Helena Parish, Louisiana (USA) were observed to have broken and frayed rectrices. The condition was noted in 21% of 90 wild turkeys harvested by hunters during the springs of 1985 through 1988 from the Pearl River Wildlife Management Area. Damage to feathers ranged from mild to severe. Histologic and microbiologic study of five birds disclosed colonization and invasion of the rachis sheath and pulp by fungi of the genera Aspergillus, Curvularia, Cladosporium, Dactylella, Exophiala, Helminthosporium and Trichophyton and by Streptomyces. Sterilized normal rectrices from wild turkeys were inoculated with these organisms and subsequently developed damage that was histologically compatible with field cases. The condition was diagnosed as a multiple etiology mycosis. Successful colonization and invasion of experimentally inoculated feathers required addition of moisture and elevation of relative humidity within the cultures. The apparent high moisture requirements of the fungi suggest that late winter and early spring flooding may be a probable predisposing factor for this condition.     

Thiol-containing cross-linking agent with enhanced steric hindrance

Ricin A chain immunotoxins disulfide cross-linked with conventional, sterically unhindered reagents have unsatisfactorily short circulating life times in vivo. (Acetylthio)succinic anhydride, a thiolating reagent with partial steric hindrance of the sulfur atom, does not remedy this situation. Sulfosuccinimidyl N-[3-(acetylthio)-3-methylbutyryl]-beta- alaninate, a new cross-linker in which the carbon alpha to the sulfur is doubly methylated, creates disulfide bonds 2 orders of magnitude more resistant to reduction than unhindered disulfides. Nevertheless, this deactivated thiolating agent rapidly and reliably cross-links ricin A chain and antibodies to create immunotoxins with in vitro cytotoxicities comparable to those of 2-iminothiolane-coupled conjugates.