Order parameter with line nodes ands±-wave symmetry for the noncentrosymmetric superconductor Li2Pt3B

Mukherjee, Soumya P.,Takimoto, Tetsuya American Physical Society 2012 Physical review. B, Condensed matter and materials Vol.86 No.13

Improved Production and Properties of β-glucosidase Influenced by 2-deoxy-D-glucose in the Culture Medium of Termitomyces clypeatus

Shakuntala Ghorai,Sumana Mukherjee,Soumya Mukherjee,Suman Khowala 한국생물공학회 2011 Biotechnology and Bioprocess Engineering Vol.16 No.2

Increased production, secretion, and activity of β-glucosidase in the filamentous fungus Termitomyces clypeatus was achieved in presence of the glycosylation inhibitor 2-deoxy-D-glucose (0.05%, w/v) during submerged fermentation. Enzyme activity increased to 163 U/mL by adding mannose (2 mg/mL) to the medium. Such a high enzyme activity has not been achieved without mutation or genetic manipulation. The K_m and V_max of the enzyme in culture medium were determined to be 0.092 mM and 35.54 U/mg, respectively, with p-nitrophenyl β-D-glucopyranoside as substrate, confirming its high catalytic activity. The enzyme displayed optimum activity at pH 5.4 and 45°C. The enzyme was fairly stable between acidic to alkaline pH and retained about 75 ~ 65% residual activities between pH 4 and 10.6 and demonstrated full activity at 45ºC for 3 days. The enzyme was also stable in the presence of Zn^2+ and Mg^2+ and 80% of the residual activity was observed in the presence of Mn^2+, Ca^2+, K^+, Cu^2+,EDTA, and sodium azide. Around 70% of the activity was retained in the presence of 2 M guanidium HCl and 3 M urea, whereas the activity was 5 and 2 times higher in the presence of 4 mM beta-mercaptoethanol and 50 mM DTT,respectively. The enzyme obtained from the culture filtrate showed potential cellulose saccharifying ability which increased further when supplemented with commercial cellulase. Thus, this enzyme could be used without any additional downstream processing for commercial cellulase preparation and production of bioethanol or for other biotechnological applications.

In situ Reversible Aggregation of Extracellular Cellobiase in the Filamentous Fungus Termitomyces clypeatus

Samudra Prosad Banik,Swagata Pal,Shakuntala Ghorai,Sudeshna Chowdhury,Rajib Majumder,Soumya Mukherjee,Suman Khowala 한국생물공학회 2012 Biotechnology and Bioprocess Engineering Vol.17 No.5

Cellobiase (E.C. 3.2.1.21), is a widely exploited industrial glycosidase with a major role in biofuel industry. Its stability and shelf life are major bottlenecks in achieving a superior formulation for industry. In the filamentous fungus Termitomyces clypeatus, the enzyme is secreted in a co-aggregated form with sucrase; the separation of this co-aggregation results in substantial loss of the enzyme’s activity. The aim of the present study was to examine the mode of aggregation of the secreted cellobiase-sucrase coaggregate and its role in the stabilization of cellobiase. Transmission electron microscopy and dynamic light scattering of purified co-aggregates revealed reversible, concentration driven self-aggregation of the extracellular enzymes to form larger entities. However, the intracellular enzyme aggregates were rigid,non-interacting, and possessed a higher percentage of disulphide bonds. Circular dichroic spectra of the two coaggregates indicated no significant difference in secondary structures. Self-association increased the stability of extracellular aggregates towards heat by 1.5 fold, SDS by 4 ~ 7 fold, and chaotropic agents, by 1.5 ~ 2 fold, than the intracellular counterpart. The Km of extracellular aggregate varied between 0.29 and 0.45 mM as a result of spontaneous aggregation and disaggregation, whereas that of intracellular aggregate was 0.22 mM irrespective of its concentration status. In situ detection of cellobiase in native PAGE revealed two activity bands of the extracellular enzyme, which indicated a minimum of two active dissociated aggregate species, as compared to a single band for the intracellular enzyme. These studies are believed to improve the understanding of aggregation of the fungal glycosidases, which remains to be a blackbox, to increase the efficacy of these enzymes. Cellobiase (E.C. 3.2.1.21), is a widely exploited industrial glycosidase with a major role in biofuel industry. Its stability and shelf life are major bottlenecks in achieving a superior formulation for industry. In the filamentous fungus Termitomyces clypeatus, the enzyme is secreted in a co-aggregated form with sucrase; the separation of this co-aggregation results in substantial loss of the enzyme’s activity. The aim of the present study was to examine the mode of aggregation of the secreted cellobiase-sucrase coaggregate and its role in the stabilization of cellobiase. Transmission electron microscopy and dynamic light scattering of purified co-aggregates revealed reversible, concentration driven self-aggregation of the extracellular enzymes to form larger entities. However, the intracellular enzyme aggregates were rigid,non-interacting, and possessed a higher percentage of disulphide bonds. Circular dichroic spectra of the two coaggregates indicated no significant difference in secondary structures. Self-association increased the stability of extracellular aggregates towards heat by 1.5 fold, SDS by 4 ~ 7 fold, and chaotropic agents, by 1.5 ~ 2 fold, than the intracellular counterpart. The Km of extracellular aggregate varied between 0.29 and 0.45 mM as a result of spontaneous aggregation and disaggregation, whereas that of intracellular aggregate was 0.22 mM irrespective of its concentration status. In situ detection of cellobiase in native PAGE revealed two activity bands of the extracellular enzyme, which indicated a minimum of two active dissociated aggregate species, as compared to a single band for the intracellular enzyme. These studies are believed to improve the understanding of aggregation of the fungal glycosidases, which remains to be a blackbox, to increase the efficacy of these enzymes.

Transcriptome profiling uncovers the involvement of CmXyn1, a glycosyl hydrolase 11, in Cochliobolus miyabeanus pathogenicity

Lee Gi Hyun,Yoo Ju Soon,Oh Ha-Ram,Min Cheol Woo,Jang Jeong Woo,Mukherjee Soumya,Jung Ki-Hong,Kim Yu-Jin,Wang Yiming,Gupta Ravi,Kim Sun Tae 한국응용생명화학회 2023 Applied Biological Chemistry (Appl Biol Chem) Vol.66 No.-

Necrotrophic pathogen Cochliobolus miyabeanus (C. miyabeanus) causes rice brown leaf spot disease and drastically affects the yield and quality of rice grains. However, the molecular mechanism of rice-C. miyabeanus remains poorly understood due to the limited research conducted on this pathosystem. To elucidate the molecular mechanism of rice-C. miyabeanus, a transcriptome analysis was conducted from in vitro and in planta grown C. miyabeanus. This analysis led to the identification of a total of 24,060 genes of which 426 in vitro and 57 in planta expressed genes were predicted to encode for secretory proteins. As these 57 genes were specifically expressed in planta and were predicted to be secretory in nature, these were consider as putative effectors, highlighting their possible roles in the fungal pathogenicity. Notably, among these putative effectors, CmXyn1 which encodes a glycosyl hydrolase 11 displayed the highest expression level under in planta conditions and was thus selected for further functional characterization. Interestingly, the extracellular expression of CmXyn1 transiently induced cell death in Nicotiana benthamiana leaves, while intracellular expression was comparatively lesser effective. In addition, transcriptome analysis on rice leaves during C. miyabeanus infection and comparing it to the rice leaf transcriptome data obtained during hemibiotrophic pathogen Magnaporthe oryzae infection led to the discovery of 18 receptors/receptor-like kinases that were commonly expressed in response to both pathogens, indicating their key roles in rice defense response. Taken together, our findings provide new insights into rice-C. miyabeanus interaction as well as the unique and common defense responses of rice against hemibiotroph and necrotroph model systems.