Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7
ABSTRACT We selected for spore-forming psychrophilic bacteria able to use lactose as a carbon source and one isolate, designated Paenibacillus sp. strain C7, that was phylogenetically related to, but distinct from both Paenibacillus macquariensis and Paenibacillus antarcticus . Some Escherichia coli...
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2005
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Online Access: | http://dx.doi.org/10.1128/aem.71.8.4225-4232.2005 https://journals.asm.org/doi/pdf/10.1128/AEM.71.8.4225-4232.2005 |
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crasmicro:10.1128/aem.71.8.4225-4232.2005 2024-06-23T07:46:17+00:00 Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7 Shipkowski, Stephanie Brenchley, Jean E. 2005 http://dx.doi.org/10.1128/aem.71.8.4225-4232.2005 https://journals.asm.org/doi/pdf/10.1128/AEM.71.8.4225-4232.2005 en eng American Society for Microbiology https://journals.asm.org/non-commercial-tdm-license Applied and Environmental Microbiology volume 71, issue 8, page 4225-4232 ISSN 0099-2240 1098-5336 journal-article 2005 crasmicro https://doi.org/10.1128/aem.71.8.4225-4232.2005 2024-06-10T04:07:18Z ABSTRACT We selected for spore-forming psychrophilic bacteria able to use lactose as a carbon source and one isolate, designated Paenibacillus sp. strain C7, that was phylogenetically related to, but distinct from both Paenibacillus macquariensis and Paenibacillus antarcticus . Some Escherichia coli transformants obtained with genomic DNA from this isolate hydrolyzed X-Gal (5-bromo-4-chloro-3-indoyl-β- d -galactopyranoside) only below 30°C, an indication of cold-active β-galactosidase activity. Sequencing of the cloned insert revealed an open reading frame encoding a 756-amino acid protein that, rather than belonging to a family typically known for β-galactosidase activity, belonged to glycoside hydrolase family 3, a family of β-glucosidases. Because of this unusual placement, the recombinant enzyme (BglY) was purified and characterized. Consistent with its classification, the enzyme had seven times greater activity with the glucoside substrate ONPGlu ( o -nitrophenyl-β- d -glucopyranoside) than with the galactoside substrate ONPGal ( o -nitrophenyl-β- d -galactopyranoside). In addition, the enzyme had, with ONPGlu, a thermal optimum around 30 to 35°C, activity over a broad pH range (5.5 to 10.9), and an especially low K m (<0.003 mM). Further examination of substrate preference showed that the BglY enzyme also hydrolyzed other aryl-β-glucosides such as helicin, MUG (4-methylumbelliferyl-β- d -glucopyranoside), esculin, indoxyl-β- d -glucoside (a natural indigo precursor), and salicin, but had no activity with glucosidic disaccharides or lactose. These characteristics and substrate preferences make the BglY enzyme unique among the family 3 β-glucosidases. The hydrolysis of a variety of aryl-β-glucosides suggests that the enzyme may allow the organism to use these substrates in the environment and that its low K m on indoxyl-β- d -glucoside may make it useful for producing indigo. Article in Journal/Newspaper Antarc* antarcticus ASM Journals (American Society for Microbiology) Applied and Environmental Microbiology 71 8 4225 4232 |
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ASM Journals (American Society for Microbiology) |
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crasmicro |
language |
English |
description |
ABSTRACT We selected for spore-forming psychrophilic bacteria able to use lactose as a carbon source and one isolate, designated Paenibacillus sp. strain C7, that was phylogenetically related to, but distinct from both Paenibacillus macquariensis and Paenibacillus antarcticus . Some Escherichia coli transformants obtained with genomic DNA from this isolate hydrolyzed X-Gal (5-bromo-4-chloro-3-indoyl-β- d -galactopyranoside) only below 30°C, an indication of cold-active β-galactosidase activity. Sequencing of the cloned insert revealed an open reading frame encoding a 756-amino acid protein that, rather than belonging to a family typically known for β-galactosidase activity, belonged to glycoside hydrolase family 3, a family of β-glucosidases. Because of this unusual placement, the recombinant enzyme (BglY) was purified and characterized. Consistent with its classification, the enzyme had seven times greater activity with the glucoside substrate ONPGlu ( o -nitrophenyl-β- d -glucopyranoside) than with the galactoside substrate ONPGal ( o -nitrophenyl-β- d -galactopyranoside). In addition, the enzyme had, with ONPGlu, a thermal optimum around 30 to 35°C, activity over a broad pH range (5.5 to 10.9), and an especially low K m (<0.003 mM). Further examination of substrate preference showed that the BglY enzyme also hydrolyzed other aryl-β-glucosides such as helicin, MUG (4-methylumbelliferyl-β- d -glucopyranoside), esculin, indoxyl-β- d -glucoside (a natural indigo precursor), and salicin, but had no activity with glucosidic disaccharides or lactose. These characteristics and substrate preferences make the BglY enzyme unique among the family 3 β-glucosidases. The hydrolysis of a variety of aryl-β-glucosides suggests that the enzyme may allow the organism to use these substrates in the environment and that its low K m on indoxyl-β- d -glucoside may make it useful for producing indigo. |
format |
Article in Journal/Newspaper |
author |
Shipkowski, Stephanie Brenchley, Jean E. |
spellingShingle |
Shipkowski, Stephanie Brenchley, Jean E. Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7 |
author_facet |
Shipkowski, Stephanie Brenchley, Jean E. |
author_sort |
Shipkowski, Stephanie |
title |
Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7 |
title_short |
Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7 |
title_full |
Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7 |
title_fullStr |
Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7 |
title_full_unstemmed |
Characterization of an Unusual Cold-Active β-Glucosidase Belonging to Family 3 of the Glycoside Hydrolases from the Psychrophilic Isolate Paenibacillus sp. Strain C7 |
title_sort |
characterization of an unusual cold-active β-glucosidase belonging to family 3 of the glycoside hydrolases from the psychrophilic isolate paenibacillus sp. strain c7 |
publisher |
American Society for Microbiology |
publishDate |
2005 |
url |
http://dx.doi.org/10.1128/aem.71.8.4225-4232.2005 https://journals.asm.org/doi/pdf/10.1128/AEM.71.8.4225-4232.2005 |
genre |
Antarc* antarcticus |
genre_facet |
Antarc* antarcticus |
op_source |
Applied and Environmental Microbiology volume 71, issue 8, page 4225-4232 ISSN 0099-2240 1098-5336 |
op_rights |
https://journals.asm.org/non-commercial-tdm-license |
op_doi |
https://doi.org/10.1128/aem.71.8.4225-4232.2005 |
container_title |
Applied and Environmental Microbiology |
container_volume |
71 |
container_issue |
8 |
container_start_page |
4225 |
op_container_end_page |
4232 |
_version_ |
1802644993430519808 |