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In ruminants, the bacterial community in the gastrointestinal tract (GIT) has an essential role in healthy growth. Examining the bacterial composition in the GIT between growth-retarded and normal yaks could improve our understanding of the role of microorganisms in yaks with growth retardation. In this study, eight male yaks with growth retardation were used as the growth-retarded yak (GRY) group, and another eight male growth normal yaks (GNYs) with the same breed and age were used as the GNY group. We compared the bacterial community in the rumen, duodenum, jejunum, ileum, cecum, and colon between GRY and GNY groups based on the 16S ribosomal RNA gene sequencing. Alpha-diversity revealed that the Shannon index in the duodenum and ileum of the GNY group was higher (P < 0.05) than that of the GRY group. However, the opposite trend was found in the jejunum and cecum. The principal coordinates analysis (PCoA) showed that the bacterial structure in all segments of GIT differed from each other between two groups. In the rumen, the relative abundances of Ruminococcaceae NK4A214 group, Ruminococcaceae UCG-014, and Treponema 2 were higher (P < 0.05) in the GNY group as compared with the GRY group. However, the Christensenellaceae R-7 group exhibited an opposite trend. In the jejunum, compared with the GNY group, the unclassified Chitinophagaceae was enriched significantly (P < 0.05) in the GRY group. However, the unclassified Peptostreptococcaceae, Christensenellaceae R-7 group, and Lachnospiraceae NK3A20 group were enriched (P < 0.05) in the GNY group. In the ileum, the relative abundances of the Rikenellaceae RC9 gut group and Prevotellaceae UCG-004 were higher (P < 0.05) in the GNY group than those in the GRY group. In the cecum, the GNY group showed a higher (P < 0.05) relative abundance of Prevotellaceae UCG-003 as compared with the GRY group. In the colon, the relative abundances of Treponema 2 and unclassified Lachnospiraceae were slightly higher (0.05 < P < 0.10) in the GNY group than those in the GRY group. Overall, these results improve our knowledge about the bacterial composition in the GIT of growth-retarded and normal yaks, and regulating the bacterial community may be an effective solution to promote the compensatory growth of GRYs.

The ocean harbors a variety of bacteria that contain huge protease resources and offer a great potential for industrial and biotechnological applications. Here, we isolated a protease-secreting bacterium Vibrio pomeroyi strain 12613 from Atlantic seawater and purified a protease VP9 from strain 12613. VP9 was identified as a metalloprotease of the M4 family. VP9 could hydrolyze casein and gelatin but not elastin and collagen. With gelatin as the substrate, VP9 showed the highest activity at 40°C and pH 6.0–8.0. It was stable at temperatures of 50°C and less and in the range of pH 5.0–11.0. VP9 also had good tolerance to NaCl, non-ionic detergents, and organic solvent methanol. Unlike other M4 metalloproteases, VP9 has distinct collagen-swelling ability, and its collagen-swelling effect was concentration dependent. The relative expansion volume of collagen increased by approximately eightfold after treatment with 10 μM VP9 at 37°C for 12 h. The collagen-swelling mechanism of VP9 on bovine-insoluble type I collagen was further studied. Atomic force microscopy observation and biochemical analyses showed that VP9 can degrade proteoglycans in collagen fibers, resulting in the release of collagen fibrils from collagen fibers and the swelling of the latter. In addition, VP9 can degrade glycoproteins, a non-collagenous constituent interacting with collagen in the skin. The characteristics of VP9, such as sufficient specificity toward proteoglycans and glycoproteins but no activity toward collagen, suggest its promising potential in the unhairing and fiber-opening processing in leather industry.

Bacterial extracellular proteases are important for bacterial nutrition and marine sedimentary organic nitrogen degradation. However, only a few proteases from marine sedimentary bacteria have been characterized. Some subtilases have a protease-associated (PA) domain inserted in the catalytic domain. Although structural analysis and deletion mutation suggests that the PA domain in subtilases is involved in substrate binding, direct evidence to support this function is still absent. Here, a protease, P57, secreted by Photobacterium sp. A5-7 isolated from marine sediment was characterized. P57 could hydrolyze casein, gelatin and collagen. It showed the highest activity at 40°C and pH 8.0. P57 is a new subtilase, with 63% sequence identity to the closest characterized protease. Mature P57 contains a catalytic domain and an inserted PA domain. The recombinant PA domain from P57 was shown to have collagen-binding ability, and Phe349 and Tyr432 were revealed to be key residues for collagen binding in the PA domain. This study first shows direct evidence that the PA domain of a subtilase can bind substrate, which provides a better understanding of the function of the PA domain of subtilases and bacterial extracellular proteases from marine sediment.