Accumulation of environmental chitin in the lungs can lead to pulmonary fibrosis, characterized by inflammatory infiltration and fibrosis in acidic chitinase (Chia)-deficient mice. Transgenic expression of Chia in these mice ameliorated the symptoms, indicating the potential of enzyme supplementation as a promising therapeutic strategy for related lung diseases. This study focuses on utilizing hyperactivated human Chia, which exhibits low activity. We achieved significant activation of human Chia by incorporating nine amino acids derived from the crab-eating monkey (Macaca fascicularis) Chia, known for its robust chitin-degrading activity. The modified human Chia retained high activity across a broad pH spectrum and exhibited enhanced thermal stability. The amino acid substitutions associated with hyperactivation of human Chia activity occurred species specifically in monkey Chia. This discovery highlights the potential of hyperactivated Chia in treating pulmonary diseases resulting from chitin accumulation in human lungs.

• Klíčová slova
• acidic chitinase (Chia), amino acid substitutions, chitin, enzyme engineering, evolution, exon swapping, hyperactivation, primate lineage, treating pulmonary diseases,
• MeSH
• chitin metabolismus   MeSH
• chitinasy * genetika   metabolismus   chemie   MeSH
• koncentrace vodíkových iontů MeSH
• lidé MeSH
• myši MeSH
• stabilita enzymů MeSH
• substituce aminokyselin MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• CHIA protein, human MeSH Prohlížeč
• chitin MeSH
• chitinasy * MeSH

Placental mammals' ancestors were insectivores, suggesting that modern mammals may have inherited the ability to digest insects. Acidic chitinase (Chia) is a crucial enzyme hydrolyzing significant component of insects' exoskeleton in many species. On the other hand, herbivorous animal groups, such as cattle, have extremely low chitinase activity compared to omnivorous species, e.g., mice. The low activity of cattle Chia has been attributed to R128H mutation. The presence of either of these amino acids correlates with the feeding behavior of different bovid species with R and H determining the high and low enzymatic activity, respectively. Evolutionary analysis indicated that selective constraints were relaxed in 67 herbivorous Chia in Cetartiodactyla. Despite searching for another Chia paralog that could compensate for the reduced chitinase activity, no active paralogs were found in this order. Herbivorous animals' Chia underwent genetic alterations and evolved into a molecule with low activity due to the chitin-free diet.

• Klíčová slova
• Evolutionary biology, Molecular biology, Zoology,
• Publikační typ
• časopisecké články MeSH

Chitooligosaccharides exhibit several biomedical activities, such as inflammation and tumorigenesis reduction in mammals. The mechanism of the chitooligosaccharides' formation in vivo has been, however, poorly understood. Here we report that mouse acidic chitinase (Chia), which is widely expressed in mouse tissues, can produce chitooligosaccharides from deacetylated chitin (chitosan) at pH levels corresponding to stomach and lung tissues. Chia degraded chitin to produce N-acetyl-d-glucosamine (GlcNAc) dimers. The block-type chitosan (heterogenous deacetylation) is soluble at pH 2.0 (optimal condition for mouse Chia) and was degraded into chitooligosaccharides with various sizes ranging from di- to nonamers. The random-type chitosan (homogenous deacetylation) is soluble in water that enables us to examine its degradation at pH 2.0, 5.0, and 7.0. Incubation of these substrates with Chia resulted in the more efficient production of chitooligosaccharides with more variable sizes was from random-type chitosan than from the block-type form of the molecule. The data presented here indicate that Chia digests chitosan acquired by homogenous deacetylation of chitin in vitro and in vivo. The degradation products may then influence different physiological or pathological processes. Our results also suggest that bioactive chitooligosaccharides can be obtained conveniently using homogenously deacetylated chitosan and Chia for various biomedical applications.

• Klíčová slova
• FACE method, acidic chitinase, block-type chitosan, chitin, chitooligosaccharides, random-type chitosan,
• MeSH
• chitinasy chemie   metabolismus   MeSH
• chitosan chemie   metabolismus   MeSH
• difrakce rentgenového záření MeSH
• hydrolýza MeSH
• koncentrace vodíkových iontů * MeSH
• myši MeSH
• oligosacharidy chemie   metabolismus   MeSH
• orgánová specificita MeSH
• plíce metabolismus   MeSH
• substrátová specifita MeSH
• žaludek metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• chitinasy MeSH
• chitosan MeSH
• oligochitosan MeSH Prohlížeč
• oligosacharidy MeSH

Chitin is a polymer of N-acetyl-D-glucosamine (GlcNAc) and a main constituent of insects' exoskeleton. Insects are rich in protein with high energy conversion efficiency. Recently, we have reported that acidic chitinases (Chia) act as digestive enzymes in mouse, pig and chicken (omnivorous) but not in dog (carnivorous) and bovine (herbivorous), indicating that feeding behavior affects Chia expression levels, and determines chitin digestibility in the particular animals. Common marmoset (Callithrix jacchus) belongs to New World monkey family and provides a potential bridge between mouse models and human diseases. Common marmoset is an insectivorous nonhuman primate with unknown expression levels and enzymatic functions of the Chia homologue, CHIA. Here, we report that common marmoset highly expresses pepsin-, trypsin- and chymotrypsin-resistant CHIA in the stomach. We show that CHIA is most active at pH 2.0 and degrades chitin and mealworm shells into GlcNAc dimers under gastrointestinal conditions. Although common marmoset and crab-eating monkey (Old World monkey) have two CHIA genes in their genomes, they primarily express one gene in the stomach. Thus, this study is the first to investigate expression levels and enzymatic functions of CHIA in a New World primate, contributing to the understanding of dietary adaptation and digestion in this taxon.

• MeSH
• Callithrix metabolismus   MeSH
• chitin metabolismus   MeSH
• chitinasy * chemie   metabolismus   MeSH
• dieta MeSH
• stravovací zvyklosti psychologie   MeSH
• žaludek enzymologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• chitin MeSH
• chitinasy * MeSH

Chitin, a polymer of N-acetyl-D-glucosamine (GlcNAc), is a major structural component in chitin-containing organism including crustaceans, insects and fungi. Mammals express two chitinases, chitotriosidase (Chit1) and acidic mammalian chitinase (AMCase). Here, we report that pig AMCase is stable in the presence of other digestive proteases and functions as chitinolytic enzyme under the gastrointestinal conditions. Quantification of chitinases expression in pig tissues using quantitative real-time PCR showed that Chit1 mRNA was highly expressed in eyes, whereas the AMCase mRNA was predominantly expressed in stomach at even higher levels than the housekeeping genes. AMCase purified from pig stomach has highest activity at pH of around 2-4 and remains active at up to pH 7.0. It was resistant to robust proteolytic activities of pepsin at pH 2.0 and trypsin and chymotrypsin at pH 7.6. AMCase degraded polymeric chitin substrates including mealworm shells to GlcNAc dimers. Furthermore, we visualized chitin digestion of fly wings by endogenous AMCase and pepsin in stomach extract. Thus, pig AMCase can function as a protease resistant chitin digestive enzyme at broad pH range present in stomach as well as in the intestine. These results indicate that chitin-containing organisms may be a sustainable feed ingredient in pig diet.

• MeSH
• chitin metabolismus   MeSH
• chitinasy genetika   izolace a purifikace   metabolismus   MeSH
• chymotrypsin metabolismus   MeSH
• dieta * MeSH
• Drosophila chemie   MeSH
• endopeptidasy metabolismus   MeSH
• gastrointestinální trakt metabolismus   MeSH
• křídla zvířecí chemie   MeSH
• messenger RNA genetika   metabolismus   MeSH
• orgánová specificita MeSH
• pepsinogen A metabolismus   MeSH
• prasata genetika   MeSH
• rozpustnost MeSH
• substrátová specifita MeSH
• Tenebrio MeSH
• tkáňové extrakty MeSH
• trypsin metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chitin MeSH
• chitinasy MeSH
• chymotrypsin MeSH
• endopeptidasy MeSH
• messenger RNA MeSH
• pepsinogen A MeSH
• tkáňové extrakty MeSH
• trypsin MeSH

Chitin, a polymer of N-acetyl-D-glucosamine (GlcNAc), functions as a major structural component in crustaceans, insects and fungi and is the second most abundant polysaccharide in the nature. Although these chitin-containing organisms have been suggested as novel animal feed resources, chitin has long been considered as indigestible fibers in the animal body. Recently, we reported that acidic chitinase (Chia) is a protease-resistant major glycosidase in mouse gastrointestinal tract (GIT) and that it digests chitin in the mouse stomach. However, the physiological role of Chia in other animals including poultry remains unknown. Here, we report that Chia can function as a digestive enzyme that breaks down chitin-containing organisms in chicken GIT. Chia mRNA is predominantly expressed in the glandular stomach tissue in normal chicken. We also show that chicken Chia has a robust chitinolytic activity at pH 2.0 and is highly resistant to proteolysis by pepsin and trypsin/chymotrypsin under conditions mimicking GIT. Chia degraded shells of mealworm larvae in the presence of digestive proteases and produced (GlcNAc)2. Thus, functional similarity of chicken Chia with the mouse enzyme suggests that chitin-containing organisms can be used for alternative poultry diets not only as whole edible resources but also as enhancers of their nutritional value.

• MeSH
• chitin metabolismus   MeSH
• chitinasy metabolismus   MeSH
• koncentrace vodíkových iontů MeSH
• kur domácí metabolismus   MeSH
• proteasy MeSH
• střeva enzymologie   MeSH
• Tenebrio chemie   MeSH
• trávení * MeSH
• žaludek enzymologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chitin MeSH
• chitinasy MeSH
• proteasy MeSH