ABSTRACT:The in vitro biodegradation of Bombyx mori silk fibroin was studied by incubating fibers and films with proteolytic enzymes (collagenase type F, ␣-chymotrypsin type I-S, protease type XXI), for times ranging from 1 to 17 days. The changes in sample weight and degree of polymerization of silk fibers exposed to proteolytic attack were negligible. However, tensile properties were significantly affected, as shown by the drop of strength and elongation as a function of the degradation time. Upon incubation with proteolytic enzymes, silk films exhibited a noticeable decrease of sample weight and degree of polymerization, the extent of which depended on the type of enzyme, on the enzyme-to-substrate ratio, and on the degradation time. Protease was more aggressive than ␣-chymotrypsin or collagenase. Film fragments resistant to enzymatic degradation were enriched in glycine and alanine. FT-IR measurements showed that the degree of crystallinity of biodegraded films increased. Soluble degradation products of silk films consisted of a range of peptides widely differing in size, deriving from the amorphous sequences of the silk fibroin chains. Biodegraded fibers showed an increase of surface roughness, while films displayed surface cracks and cavities with internal voids separated by fiber-like elements.

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[NiFe] hydrogenase maturation proteins HypC, HypD, and HypE catalyze the insertion and cyanation of the iron center of [NiFe] hydrogenases by an unknown mechanism. We have determined the crystal structures of HypC, HypD, and HypE from Thermococcus kodakaraensis KOD1 at 1.8 A, 2.07 A, and 1.55 A resolution, respectively. The structure of HypD reveals its probable iron binding and active sites for cyanation. An extended conformation of each conserved motif of HypC and HypE allows the essential cysteine residues of both proteins to interact with the active site of HypD. Furthermore, the C-terminal tail of HypE is shown to exist in an ATP-dependent dynamic equilibrium between outward and inward conformations. Unexpectedly, the [4Fe-4S] cluster environment of HypD is quite similar to that of ferredoxin:thioredoxin reductase (FTR), indicating the existence of a redox cascade similar to the FTR system. These results suggest a cyanation reaction mechanism via unique thiol redox signaling in the HypCDE complex.

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Sulfurovum aggregans sp. nov., a hydrogenoxidizing, thiosulfate-reducing chemolithoautotroph within the Epsilonproteobacteria isolated from a deep-sea hydrothermal vent chimney, and an emended description of the genus Sulfurovum , was isolated from a deep-sea hydrothermal vent chimney at the Central Indian Ridge. The non-motile, rod-shaped cells were Gram-stain-negative and non-sporulating. Growth was observed between 15 and 37 6C (optimum 33 6C; 3.2 h doubling time) and between pH 5.4 and 8.6 (optimum pH 6.0). The isolate was a strictly anaerobic chemolithoautotroph capable of using molecular hydrogen as the sole energy source and carbon dioxide as the sole carbon source. The G+C content of the genomic DNA was 42.6 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the novel isolate belonged to the genus Sulfurovum and was closely related to Sulfurovum sp. NBC37-1 and Sulfurovum lithotrophicum 42BK T (95.6 and 95.4 % similarity, respectively). DNA-DNA hybridization demonstrated that the novel isolate could be differentiated genotypically from Sulfurovum sp. NBC37-1 and Sulfurovum lithotrophicum. On the basis of the molecular and physiological traits of the new isolate, the name Sulfurovum aggregans sp. nov. is proposed, with the type strain Monchim33 T (5JCM 19824Deep-sea hydrothermal vents represent one of the most productive marine ecosystems. The ecosystem is supported primarily by microbial chemosynthesis (Jannasch, 1985). Deep-sea vent chemolithoautotrophs are able to use a range of reductive substrates as energy sources (Campbell et al., 2006;Fisher et al., 2007;Sievert & Vetriani, 2012). Molecular hydrogen is one of the most abundant reducing gas components in vent fluids. A diversity of hydrogen-oxidizing, chemolithoautotrophic, thermophilic and hyperthermophilic genera have been isolated from deep-sea hydrothermal fields (Sievert & Vetriani, 2012). In addition, recent research indicated that H 2 could be an important energy source for symbiotic psychrophilic to mesophilic bacteria (Petersen et al., 2011). Various mesophilic, hydrogen-oxidizing Epsilonproteobacteria have been isolated from deep-sea vents, including Sulfurimonas paralvinellae GO25 T , Sulfurovum sp. NBC37-1 (Nakagawa et al., 2007), Thioreductor micantisoli BKB25Ts-Y T (Nakagawa et al., 2005a) and 'Thiofractor thiocaminus' 496Chim (Makita et al., 2012). In particular, members of the genus Sulfurovum are frequently found as endo-and/or epi-symbionts of various vent animals in global deep-sea hydrothermal fields (Haddad et al., 1995;Polz & Cavanaugh, 1995;Cary et al., 1997;Goffredi et al., 2004; Suzuki et al., 2005; Urakawa et al., 2005;Watsuji et al., 2010). However, the description of the genus Sulfurovum is based on just two strains originated from the 3Present address: