Identification of essential amino acid residues for catalytic activity and thermostability of novel chitosanase by site-directed mutagenesis

2001 ◽  
Vol 56 (1-2) ◽  
pp. 173-180 ◽  
Author(s):  
H.-G. Yoon ◽  
H.-Y. Kim ◽  
Y.-H. Lim ◽  
H.-K. Kim ◽  
D.-H. Shin ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Amino Acid ◽  
Essential Amino Acid ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Amino Acid Residues

Analysis of essential carboxylic amino acid residues for catalytic activity of fungal chitosanases by site-directed mutagenesis

2005 ◽  
Vol 100 (5) ◽  
pp. 545-550 ◽  
Author(s):  
Makoto Shimosaka ◽  
Kazuaki Sato ◽  
Naohide Nishiwaki ◽  
Takashi Miyazawa ◽  
Mitsuo Okazaki
Keyword(s):  
Catalytic Activity ◽  
Amino Acid ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Amino Acid Residues

scholarly journals Riboflavin Synthase ofEscherichia coli

2001 ◽  
Vol 276 (15) ◽  
pp. 11524-11530 ◽  
Author(s):  
Boris Illarionov ◽  
Kristina Kemter ◽  
Sabine Eberhardt ◽  
Gerald Richter ◽  
Mark Cushman ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Catalytic Activity ◽  
Amino Acid ◽  
Binding Sites ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Sequence Motif ◽  
Amino Acid Residues ◽  
Terminal Sequence ◽  
Substrate Binding Sites

Conserved amino acid residues of riboflavin synthase fromEscherichia coliwere modified by site-directed mutagenesis. Replacement or deletion of phenylalanine 2 afforded catalytically inactive proteins. S41A and H102Q mutants had substantially reduced reaction velocities. Replacements of various other conserved polar residues had little impact on catalytic activity.19F NMR protein perturbation experiments using a fluorinated intermediate analog suggest that the N-terminal sequence motif MFTG is part of one of the substrate-binding sites of the protein.

Chitosanase from Streptomyces sp. strain N174: a comparative review of its structure and function

1997 ◽  
Vol 75 (6) ◽  
pp. 687-696 ◽  
Author(s):  
Tamo Fukamizo ◽  
Ryszard Brzezinski
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Substrate Binding ◽  
Structure And Function ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Amino Acid Residues ◽  
Streptomyces Sp ◽  
Binding Cleft ◽  
And Function

Novel information on the structure and function of chitosanase, which hydrolyzes the beta -1,4-glycosidic linkage of chitosan, has accumulated in recent years. The cloning of the chitosanase gene from Streptomyces sp. strain N174 and the establishment of an efficient expression system using Streptomyces lividans TK24 have contributed to these advances. Amino acid sequence comparisons of the chitosanases that have been sequenced to date revealed a significant homology in the N-terminal module. From energy minimization based on the X-ray crystal structure of Streptomyces sp. strain N174 chitosanase, the substrate binding cleft of this enzyme was estimated to be composed of six monosaccharide binding subsites. The hydrolytic reaction takes place at the center of the binding cleft with an inverting mechanism. Site-directed mutagenesis of the carboxylic amino acid residues that are conserved revealed that Glu-22 and Asp-40 are the catalytic residues. The tryptophan residues in the chitosanase do not participate directly in the substrate binding but stabilize the protein structure by interacting with hydrophobic and carboxylic side chains of the other amino acid residues. Structural and functional similarities were found between chitosanase, barley chitinase, bacteriophage T4 lysozyme, and goose egg white lysozyme, even though these proteins share no sequence similarities. This information can be helpful for the design of new chitinolytic enzymes that can be applied to carbohydrate engineering, biological control of phytopathogens, and other fields including chitinous polysaccharide degradation. Key words: chitosanase, amino acid sequence, overexpression system, reaction mechanism, site-directed mutagenesis.

Identification of Amino Acid Residues Underlying the Antiport Mechanism of the Mitochondrial Carnitine/Acylcarnitine Carrier by Site-Directed Mutagenesis and Chemical Labeling

Biochemistry ◽  
2014 ◽  
Vol 53 (44) ◽  
pp. 6924-6933 ◽  
Author(s):  
Nicola Giangregorio ◽  
Lara Console ◽  
Annamaria Tonazzi ◽  
Ferdinando Palmieri ◽  
Cesare Indiveri
Keyword(s):  
Amino Acid ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Amino Acid Residues ◽  
Chemical Labeling

Analysis of essential amino acid residues for catalytic activity of cis-epoxysuccinate hydrolase from Bordetella sp. BK-52

2013 ◽  
Vol 98 (4) ◽  
pp. 1641-1649 ◽  
Author(s):  
Wenna Bao ◽  
Haifeng Pan ◽  
Zhenhong Zhang ◽  
Yongqing Cheng ◽  
Zhipeng Xie ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Amino Acid ◽  
Essential Amino Acid ◽  
Amino Acid Residues
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