Improvement of the catalytic performance of chitosanase Csn-PD from Paenibacillus dendritiformis by semi-rational design.

Chitooligosaccharides (COS) possess versatile functional properties that have found extensive applications across various fields. Chitosanase can specifically hydrolyze ß-1,4 glycosidic bonds in chitosan to produce COS. In this study, Csn-PD, a glycoside hydrolase family 46 chitosanase from Paenibacillus dendritiformis, which produces (GlcN)2 as its main product, was rationally redesigned aiming to improve its catalytic performance. Based on the results of molecular docking analysis and multiple sequence alignment, four amino acid residues in Csn-PD (I101, T120, T220, and Y259) were pinpointed for targeted mutations. Beneficial mutations in terms of enhanced catalytic activity were then combined by site-directed mutagenesis. Notably, the most promising variant, Csn-PDT6 (Csn-PD I101M/T120E/T220G), exhibited an impressive eight-fold surge in activity compared to the wild-type Csn-PD. This heightened enzymatic activity was complemented by an enhanced pH stability profile. A compelling feature of Csn-PDT6 is its preservation of the hydrolytic product profile observed in Csn-PD. This characteristic further accentuates its candidacy for the targeted production of (GlcN)2. The success of our strategic approach is vividly illustrated by the significant improvements achieved in the catalytic performance of the chitosanase, encompassing both its activity and stability. These developments offer a valuable model that may have implications for the semi-rational design of other enzymes.

The Intestinal Bacterial Community and Functional Potential of Litopenaeus vannamei in the Coastal Areas of China.

Intestinal bacteria are crucial for the healthy aquaculture of Litopenaeus vannamei, and the coastal areas of China are important areas for concentrated L. vannamei cultivation. In this study, we evaluated different compositions and structures, key roles, and functional potentials of the intestinal bacterial community of L. vannamei shrimp collected in 12 Chinese coastal cities and investigated the correlation between the intestinal bacteria and functional potentials. The dominant bacteria in the shrimp intestines included Proteobacteria, Bacteroidetes, Tenericutes, Firmicutes, and Actinobacteria, and the main potential functions were metabolism, genetic information processing, and environmental information processing. Although the composition and structure of the intestinal bacterial community, potential pathogenic bacteria, and spoilage organisms varied from region to region, the functional potentials were homeostatic and significantly (p < 0.05) correlated with intestinal bacteria (at the family level) to different degrees. The correlation between intestinal bacteria and functional potentials further suggested that L. vannamei had sufficient functional redundancy to maintain its own health. These findings help us understand differences among the intestinal bacterial communities of L. vannamei cultivated in different regions and provide a basis for the disease management and healthy aquaculture of L. vannamei.

Cloning, expression and comparison of the properties of Scy p 9, a Scylla paramamosain allergen.

This study investigated the properties of Scy p 9 in mud crab (Scylla paramamosain). The gene sequence of filamin C obtained from crabs, which was denoted as Scy p 9, contains a 2544 bp open reading frame and encodes 848 amino acid residues. Recombinant Scy p 9 (rScy p 9) is expressed in Escherichia coli, which exhibits tertiary structure changes, and the IgE binding activity of rScy p 9 is higher than that of native Scy p 9 (nScy p 9). Moreover, this study explored the possibility of the presence and cross-reactivity of filamin C in 8 shellfish. IgE-specific binding to nScy p 9 and rScy p 9 in patients allergic to shellfish revealed that rScy p 9 was more sensitive than nScy p 9. The gene sequence of filamin C fills in the blank in shellfish. This study contributes to the understanding of the properties of Scy p 9, and the results indicate that rScy p 9 can be used as a candidate for component-resolved diagnosis in shellfish.

MicroRNA-100 functions as a tumor suppressor by inhibiting Lgr5 expression in colon cancer cells.

Previous studies have demonstrated that microRNAs (miRNAs), a class of single­stranded RNA molecules that are 18­27 nucleotides in length, serve a critical function in tumorigenesis, including in the development of colon cancer. In the current study, miR­100 levels were demonstrated to be reduced in colon cancer tissues compared with the levels in matched adjacent normal tissues. Forced overexpression of miR­100 by transfection with miR­100 mimics substantially inhibited the proliferation, migration and invasion of SW480 and HCT116 cells, whereas reduced expression, resulting from transfection of antisense oligonucleotides, promoted these processes. At the molecular level, miR­100 was observed to reduce the levels of leucine­rich repeat­containing G protein­coupled receptor 5 (Lgr5), by binding to its 3'­untranslated region. As a result of this, Wnt/ß­catenin signaling was affected by fluctuations in the level of miR­100 mimics or antisense. Collectively, the results of the current study elucidate a novel regulatory pathway involving miR­100 and Lgr5 in colon cancer cells, which may present a potential therapeutic target.