Expression and Characterization of Endochitinase C from<i>Serratia marcescens</i>BJL200 and Its Purification by a One-Step General Chitinase Purification Method

Synstad, Bjørnar; Vaaje‐Kolstad, Gustav; Cederkvist, F. Henning; Saua, Silje F.; Horn, Svein Jarle; Eijsink, Vincent G. H.; Sørlie, Morten

doi:10.1271/bbb.70594

Cited by 52 publications

(43 citation statements)

References 39 publications

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“…3 might provide a basis for the possible involvement of the C-terminal ChBD between the insoluble α-chitin substrate binding and the SmChiC enzyme, but other interactions such as the presence of FnIII and other neighboring aromatic amino acid residues could not be totally excluded. These results obtained from this study were not completely consistent with previous findings in binding studies of ChiC1 and ChiC2 of S. marcescens 2170 toward the regenerated chitin, colloidal chitin, and powdered chitin [21,22,24,25]. Many C-terminally truncated chitinases have been reported to lose their chitin-binding abilities and hydrolyzing efficiencies toward various insoluble chitin substrates [13,37,[39][40][41].…”

Section: Discussioncontrasting

confidence: 83%

Effects of C-Terminal Domain Truncation on Enzyme Properties of Serratia marcescens Chitinase C

Lin

Chen

et al. 2015

Appl Biochem Biotechnol

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A chitinase gene (SmChiC) and its two C-terminal truncated mutants, SmChiCG426 and SmChiCG330 of Serratia marcescens, were constructed and cloned by employing specific polymerase chain reaction (PCR) techniques. SmChiCG426 is derived from SmChiC molecule without its C-terminal chitin-binding domain (ChBD) while SmChiCG330 is truncated from SmChiC by its C-terminal deletion of both ChBD and fibronectin type III domain (FnIII). To study the role of the C-terminal domains of SmChiC on the enzyme properties, SmChiC, SmChiCG426, and SmChiCG330 were expressed in Escherichia coli by using the pET-20b(+) expression system. The His-tag affinity-purified SmChiC, SmChiCG426, and SmChiCG330 enzymes had a calculated molecular mass of 51, 46, and 36 kDa, respectively. Certain biochemical characterizations indicated that the enzymes had similar physicochemical properties, such as the optimum pH (5), temperature (37 °C), thermostability (50 °C), and identical hydrolyzing product (chitobiose) from both the soluble and insoluble chitin substrates. The overall catalytic efficiency k cat /K M was higher for both truncated enzymes toward the insoluble α-chitin, whereas the binding abilities toward the insoluble α-chitin substrate were reduced moderately. The fluorescence and circular dichroism (CD) spectroscopy data suggested that both mutants retained a similar folding conformation as that of the full-length SmChiC enzyme. However, a CD-monitored melting study showed that the SmChiCG330 had no obvious transition temperature, unlike the SmChiC and SmChiCG426.

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Section: Discussioncontrasting

confidence: 83%

Effects of C-Terminal Domain Truncation on Enzyme Properties of Serratia marcescens Chitinase C

Lin

Chen

et al. 2015

Appl Biochem Biotechnol

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“…44. Final purification was obtained by chitin affinity chromatography and hydrophobic interaction chromatography to isolate ChiC2.…”

Section: Methodsmentioning

confidence: 99%

Hallmarks of Processivity in Glycoside Hydrolases from Crystallographic and Computational Studies of the Serratia marcescens Chitinases

Payne

Baban

Horn

et al. 2012

Journal of Biological Chemistry

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112

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Background: Nature employs processive and nonprocessive glycoside hydrolases to degrade polysaccharides. Results: We solved the Serratia marcescens nonprocessive chitinase (ChiC2) structure and used simulation to identify dynamic hallmarks of processivity in S. marcescens chitinases. Conclusion: Dynamic metrics complement structural insights in determining processivity. Significance: Identification of hallmarks of processivity is a key step toward development of a general, molecular-level theory of glycoside hydrolase processivity.

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“…All of these chitinases retained 60% activity at alkali pH values between 7 and 11, although the highest activity was observed at pHs 8, 9, and 8.5 for ChiA, ChiB, and ChiC, respectively. The optimum pH values for the chitinases of S. marcescens were reported at 5.5 (Gal et al, 1998), 6.2 (Nawani andKapadnis, 2001), 5-6 (Brurberg et al, 1996), 5-7.5 (Lan et al, 2006), and 4-10 (Suzuki et al, 2002) for ChiA; 4-10 ( Suzuki et al, 2002;Bahar et al, 2012) and 5-6 (Brurberg et al, 1996) for ChiB; and 3-9 (Synstad et al, 2008) and4-10 (Suzuki et al, 2002) for ChiC. Researchers also reported bimodal pH distributions at around pH 4.0 and 8.0-9.0 (Okay et al, 2008).…”

Section: Discussionmentioning

confidence: 99%

“…Chitinases have been detected in a great variety of organisms, including those that contain chitin, such as insects, crustaceans, yeasts, and fungi, and also organisms that do not contain chitin, such as bacteria, higher plants, and vertebrates. Several genera of bacteria, including Serratia (Watanabe et al, 1997;Suzuki et al, 2002), Enterobacter (Chernin et al, 1995), and Aeromonas (Weisburg et al, 1991;Wu et al, 2001;Synstad et al, 2008;Gökçe et al, 2010) produce high levels of chitinolytic enzymes (Nawani and Kapadnis, 2001). Others, such as Bacillus thuringiensis, produce chitinases in small amounts although they can be genetically manipulated to improve their chitinase production (Bhushan and Hoondal, 1998;BarbozaCorona et al, 1999;Lauzon et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

Cloning and expression of chitinase A, B, and C (chiA, chiB, chiC) genes from Serratia marcescens originating from Helicoverpa armigera and determining their activities

Danismazoglu¹,

Demir²,

Sezen³

et al. 2015

Turk J Biol

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IntroductionChitin, a linear polymer of β-1,4-linked N-acetyl-Dglucosamine (GlcNAc), is one of the most abundant biopolymers in nature. It is the main component of the exoskeleton of many invertebrates such as crustaceans, insects, and spiders and a structural component of the cell walls of most fungi and algae. In insects it functions as scaffold material, supporting the cuticles of the epidermis and trachea as well as the peritrophic membrane (PM) lining the gut epithelium (Merzendorfer and Zimoch, 2003;Ruiz-Sanchez et al., 2005;Kuzu, 2008).Chitinases are glycosyl hydrolases that catalyze the hydrolytic degradation of chitin. Chitin, which is found in the structure of cuticles and peritrophic membranes, is degraded by the chitinase enzymes. The degradation decreases the feeding and defenses of the insect and weakens it. Therefore, chitinases are important enzymes that have potential for being used as biological control agents against harmful insects (Cohen, 1987;Debaditya et al., 2007;Kuzu, 2008). Chitinases have been detected in a great variety of organisms, including those that contain chitin, such as insects, crustaceans, yeasts, and fungi, and also organisms that do not contain chitin, such as bacteria, higher plants, and vertebrates. Several genera of bacteria, including Serratia (

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Expression and Characterization of Endochitinase C fromSerratia marcescensBJL200 and Its Purification by a One-Step General Chitinase Purification Method

Cited by 52 publications

References 39 publications

Effects of C-Terminal Domain Truncation on Enzyme Properties of Serratia marcescens Chitinase C

Effects of C-Terminal Domain Truncation on Enzyme Properties of Serratia marcescens Chitinase C

Hallmarks of Processivity in Glycoside Hydrolases from Crystallographic and Computational Studies of the Serratia marcescens Chitinases

Cloning and expression of chitinase A, B, and C (chiA, chiB, chiC) genes from Serratia marcescens originating from Helicoverpa armigera and determining their activities

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