Purification of two chitinases from Rhizopus oligosporus and isolation and sequencing of the encoding genes.
TLDR
Two chitinases were purified from Rhizopus oligosporus, a filamentous fungus belonging to the class Zygomycetes, and it is concluded that these chit inases are synthesized with pre- and prosequences in addition to the mature enzyme sequences and that the pro sequences are located at the C terminal.Abstract:
Two chitinases were purified from Rhizopus oligosporus, a filamentous fungus belonging to the class Zygomycetes, and designated chitinase I and chitinase II. Their N-terminal amino acid sequences were determined, and two synthetic oligonucleotide probes corresponding to these amino acid sequences were synthesized. Southern blot analyses of the total genomic DNA from R. oligosporus with these oligonucleotides as probes indicated that one of the two genes encoding these two chitinases was contained in a 2.9-kb EcoRI fragment and in a 3.6-kb HindIII fragment and that the other one was contained in a 2.9-kb EcoRI fragment and in a 11.5-kb HindIII fragment. Two DNA fragments were isolated from the phage bank of R. oligosporus genomic DNA with the synthetic oligonucleotides as probes. The restriction enzyme analyses of these fragments coincided with the Southern blot analyses described above and the amino acid sequences deduced from their nucleotide sequences contained those identical to the determined N-terminal amino acid sequences of the purified chitinases, indicating that each of these fragments contained a gene encoding chitinase (designated chi 1 and chi 2, encoding chitinase I and II, respectively). The deduced amino acid sequences of these two genes had domain structures similar to that of the published sequence of chitinase of Saccharomyces cerevisiae, except that they had an additional C-terminal domain. Furthermore, there were significant differences between the molecular weights experimentally determined with the two purified enzymes and those deduced from the nucleotide sequences for both genes. Analysis of the N- and C-terminal amino acid sequences of both chitinases and comparison of them with the amino acid sequences deduced from the nucleotide sequences revealed posttranslational processing not only at the N-terminal signal sequences but also at the C-terminal domains. It is concluded that these chitinases are synthesized with pre- and prosequences in addition to the mature enzyme sequences and that the prosequences are located at the C terminal.read more
Citations
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Chitinases of fungi and plants: their involvement in morphogenesis and host‐parasite interaction
A.S. Sahai,M.S. Manocha +1 more
TL;DR: Chitinase induction in plants is not considered solely as an antifungal resistance mechanism, but there is some circumstantial evidence to suggest a morphogenetic role despite the apparent absence of the substrate in plant cells.
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Microbial hydrolysis of polysaccharides.
TL;DR: In general, the systems produced by different microorganisms for the hydrolysis of a particular polysaccharide comprise similar enzymes from the same families, which are complex on two quite different levels.
Journal ArticleDOI
Identification of glutamic acid 204 and aspartic acid 200 in chitinase A1 of Bacillus circulans WL-12 as essential residues for chitinase activity.
TL;DR: Kinetic parameters for 4-methylumbelliferyl-N,N',N"-triacetylchitotriose hydrolysis were determined and results indicate a lysozyme-type catalytic mechanism of the chitinase.
Journal ArticleDOI
Review of fungal chitinases.
TL;DR: The present review will focus on recent advances of fungal chitinases, containing their nomenclature and assays, purification and characterization, molecular cloning and expression, family and structure, regulation, and function and application.
Journal ArticleDOI
Structural features of plant chitinases and chitin-binding proteins
TL;DR: As the currently known three‐dimensional structures of chitinases are those from barley and the rubber tree, Hevea brasiliensis, it is proposed to adopt the designation b‐type (classes I, II and IV) and h‐ type (classes III and V) chit inases, respectively.
References
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