Identification of a β-glucosidase from the Mucor circinelloides genome by peptide pattern recognition |
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| Institution: | 1. Section for Sustainable Biotechnology, Department of Biotechnology, Chemistry and Environmental Engineering, Aalborg University Copenhagen, DK-2450 Copenhagen SV, Denmark;2. Environmental Microbiology Key Laboratory of Sichuan Province, Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu, Sichuan 610041, China;3. University of the Chinese Academy of Sciences, Beijing 100049, China;1. Universidad Metropolitana de Ciencias de la Educación, Instituto de Entomología, Santiago, Chile;2. Universidad Autónoma del Estado de Morelos, Facultad de Ciencias Agropecuarias, Cuernavaca Morelos, Mexico;1. Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA, USA;2. Department of Plant Pathology, Nanjing Agricultural University, Nanjing, China;3. South Texas Center for Emerging Infectious Diseases (STCEID), Department of Biology, University of Texas at San Antonio, San Antonio, TX, USA;4. Department of Pediatrics, Louisiana State University Health Sciences Center, New Orleans, LA, USA;1. Laboratório de Química Orgânica e Biocatálise, Instituto de Química de São Carlos, Universidade de São Paulo, Av. João Dagnone, 1100, Ed. Química Ambiental, J. Santa Angelina, 13563-120, São Carlos, SP, Brazil;2. Grupo de Físico-Química Orgânica, Instituto de Química de São Carlos, Universidade de São Paulo (USP), Av. Trabalhador São Carlense, 400, Caixa Postal 780, 13560-970, São Carlos, SP, Brazil;3. Laboratório Nacional de Ciência e Tecnologia do Bioetanol (CTBE), Centro Nacional de Pesquisa em Energia e Materiais (CNPEM), Caixa Postal 6192, 13083-970, Campinas, SP, Brazil;4. Bioengineering Department, Instituto Superior Técnico, University of Lisbon, Institute for Bioengineering and Biosciences, Av. Rovisco Pais, 1049-001 Lisbon, Portugal;5. Escola de Engenharia de São Carlos (EESC), Universidade de São Paulo (USP), Caixa Postal 780, 13560-970, São Carlos, SP, Brazil;1. BRD School of Biosciences, Sardar Patel Maidan, Satellite Campus, P. Box No. 39, Sardar Patel University, Vallabh Vidyanagar, 388120 Gujarat, India;2. Sardar Patel Renewable Energy Research Institute, P. Box No. 2, Vallabh Vidyanagar, 388120 Gujarat, India, India;1. Department of Food Science and Technology, College of Light Industry and Food Engineering, Nanjing Forestry University, Nanjing 210037, China;2. Department of Biology, Lakehead University, Ontario P7B 5E1, Canada;3. Co-innovation Center for the Sustainable Forestry in Southern China, College of Forestry, Nanjing Forestry University, Nanjing 210037, China;4. Institute of Agricultural Products Processing, Shanxi Academy of Agricultural Science, Tianyuan 030031, China |
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| Abstract: | Mucor circinelloides produces plant cell wall degrading enzymes that allow it to grow on complex polysaccharides. Although the genome of M. circinelloides has been sequenced, only few plant cell wall degrading enzymes are annotated in this species. We applied peptide pattern recognition, which is a non-alignment based method for sequence analysis to map conserved sequences in glycoside hydrolase families. The conserved sequences were used to identify similar genes in the M. circinelloides genome. We found 12 different novel genes encoding members of the GH3, GH5, GH9, GH16, GH38, GH47 and GH125 families in M. circinelloides. One of the two GH3-encoding genes was predicted to encode a β-glucosidase (EC 3.2.1.21). We expressed this gene in Pichia pastoris KM71H and found that the purified recombinant protein had relative high β-glucosidase activity (1.73 U/mg) at pH5 and 50 °C. The Km and Vmax with p-nitrophenyl-β-d-glucopyranoside as substrate was 0.20 mM and 2.41 U/mg, respectively. The enzyme was not inhibited by glucose and retained 84% activity at glucose concentrations up to 140 mM. Although zygomycetes are not considered to be important degraders of lignocellulosic biomass in nature, the present finding of an active β-glucosidase in M. circinelloides demonstrates that enzymes from this group of fungi have a potential for cellulose degradation. |
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| Keywords: | Peptide pattern recognition β-Glucosidase Glycoside hydrolase 3 |
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