scholarly journals Coordinately express hemicellulolytic enzymes in Kluyveromyces marxianus to improve the saccharification and ethanol production from corncobs

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Qing Lan ◽  
Yitong Duan ◽  
Pingping Wu ◽  
Xueyin Li ◽  
Yao Yu ◽  
...  
Keyword(s):  
Ethanol Production ◽  
High Efficiency ◽  
Ethanol Yield ◽  
Stop Codon ◽  
Signal Sequence ◽  
Xylanase Activity ◽  
Foot And Mouth Disease ◽  
High Growth ◽  
High Growth Rate ◽  
Mouth Disease Virus

Abstract Background Hemicellulose acts as one factor contributing to the recalcitrance of lignocellulose that prevents cellulases to degrade the cellulose efficiently even in low quantities. Supplement of hemicellulases can enhance the performance of commercial cellulases in the enzymatic hydrolyses of lignocellulose. Kluyveromyce marxianus is an attractive yeast for cellulosic ethanol fermentation, as well as a promising host for heterologous protein production, since it has remarkable thermotolerance, high growth rate, and broad substrate spectrum etc. In this study, we attempted to coordinately express multiple hemicellulases in K.marxianus through a 2A-mediated ribosome skipping to self-cleave polyproteins, and investigated their capabilities for saccharification and ethanol production from corncobs. Results Two polycistronic genes IMPX and IMPαX were constructed to test the self-cleavage of P2A sequence from the Foot-and-Mouth Disease virus (FMDV) in K.marxianus. The IMPX gene consisted of a β-mannanase gene M330 (without the stop codon), a P2A sequence and a β-xylanase gene Xyn-CDBFV in turn. In the IMPαX gene, there was an additional α-factor signal sequence in frame with the N-terminus of Xyn-CDBFV. The extracellular β-mannanase activities of the IMPX and IMPαX strains were 21.34 and 15.50 U/mL, respectively, but the extracellular β-xylanase activity of IMPαX strain was much higher than that of the IMPX strain, which was 136.17 and 42.07 U/mL, respectively. Subsequently, two recombinant strains, the IXPαR and IMPαXPαR, were constructed to coordinately and secretorily express two xylantic enzymes, Xyn-CDBFV and β-D-xylosidase RuXyn1, or three hemicellulolytic enzymes including M330, Xyn-CDBFV and RuXyn1. In fed-batch fermentation, extracellular activities of β-xylanase and β-xylosidase in the IXPαR strain were 1664.2 and 0.90 U/mL. Similarly, the IMPαXPαR strain secreted the three enzymes, β-mannanase, β-xylanase, and β-xylosidase, with the activities of 159.8, 2210.5, and 1.25 U/mL, respectively. Hemicellulolases of both strains enhanced the yields of glucose and xylose from diluted acid pretreated (DAP) corncobs when acted synergistically with commercial cellulases. In hybrid saccharification and fermentation (HSF) of DAP corncobs, hemicellulases of the IMPαXPαR strain increased the ethanol yield by 8.7% at 144 h compared with the control. However, both ethanol and xylose yields were increased by 12.7 and 18.2%, respectively, at 120 h in HSF of aqueous ammonia pretreated (AAP) corncobs with this strain. Our results indicated that coordinate expression of hemicellulolytic enzymes in K. marxianus promoted the saccharification and ethanol production from corncobs. Conclusions The FMDV P2A sequence showed high efficiency in self-cleavage of polyproteins in K. marxianus and could be used for secretory expression of multiple enzymes in the presence of their signal sequences. The IMPαXPαR strain coexpressed three hemicellulolytic enzymes improved the saccharification and ethanol production from corncobs, and could be used as a promising strain for ethanol production from lignocelluloses.

scholarly journals Coordinately Express Hemicellulolytic Enzymes in Kluyveromycesmarxianus to Improve the Saccharification and Ethanol Production From Corncobs

2021 ◽  
Author(s):  
Qing Lan ◽  
Yitong Duan ◽  
Pingping Wu ◽  
Xueyin Li ◽  
Yao Yu ◽  
...  
Keyword(s):  
Ethanol Production ◽  
High Efficiency ◽  
Consolidated Bioprocessing ◽  
Stop Codon ◽  
Signal Sequence ◽  
Foot And Mouth Disease ◽  
High Growth ◽  
High Growth Rate ◽  
Cleavage Efficiency ◽  
Mouth Disease Virus

Abstract Background:Hemicelluloses act as one factor contributing to the recalcitrance of lignocelluloses that prevent cellulases to degrade the cellulose efficiently even in low quantities, and supplement of hemicellulases can enhance performance of commercial cellulases in the enzymatic hydrolyses of lignocellulose. K. marxianu is an attractive yeast for cellulosic ethanol fermentation, since it has remarkable thermotolerance, high growth rate, and broad substrate spectrum etc, as well as a promising host for heterologous protein production. In this study, we attempted to coordinately express multiple hemicellulases in Kluyveromyces marxianus through a 2A-mediated ribosomes skipping to self-cleave polyproteins, and investigated their capabilities for saccharification and ethanol production from corncobs.ResultsTwo polycistronic genes IMPX and IMPαX were constructed to test the self-cleavage efficiency of P2A sequence from Foot and Mouth Disease virus (FMDV) in K. marxianus. The IMPX gene consisted of a β-mannanase gene M330 (without the stop codon), a P2A sequence and a β-xylanase gene Xyn-CDBFV in turn, while in the IMPαX gene there was an additional α-factor signal sequence fused at the N-terminus of Xyn-CDBFV. The extracellular β-mannanase activities of IMPX and IMPαX strains were 21.34 and 15.50 U/mL repectively. By contrast, the IMPαX strain secreted 136.17 U/mL β-xylanase, which was much higher than that of IMPX strain, 42.07 U/mL. Based on these, two recombinant strains, the IXαR and IMPαXPαR, were constructed to coordinately and secretorily express the β-D-xylosidase RuXyn1 and Xyn-CDBFV, or three hemicellulolytic enzymes including M330, Xyn-CDBFV and RuXyn1. The IMPαX strain produced 1664.2 and 0.90 U/mL of extracellular β-xylanase and β-xylosidase, while the IMPαXPαR strain secreted 159.8, 2210.5, and 1.25 U/ml of β-mannanase, β-xylanase, and β-xylosidase in fed-batch fermentations respectively. Hemicellulolytic enzymes of these two strains enhanced the releases of both glucose and xylose from diluted acid pretreated corncobs when acted synergistically with commercial cellulases. In hybrid saccharification and fermentation (HSF) of pretreated corncobs, hemicellulases of the IMPαXPαR strain increased about 34.2% and 11.1% of ethanol productions at 144 and 216 h respectively .ConclusionsThe FMDV P2A sequence showed high efficiency in self-cleavage of polyproteins in K. marxianus, and could be used for secretory expression of multiple enzymes in present of their own signal sequences. The IMPαXPαR strain that coexpressed three hemicellulolytic enzymes could be used as a consolidated bioprocessing (CBP) strain for ethanol production from lignocelluloses.

scholarly journals Coordinately Express Hemicellulolytic Enzymes in Kluyveromyces Marxianus to Improve the Saccharification and Ethanol Production from Corncobs

2021 ◽  
Author(s):  
Qing Lan ◽  
Yitong Duan ◽  
Pingping Wu ◽  
Xueyin Li ◽  
Yao Yu ◽  
...  
Keyword(s):  
Ethanol Production ◽  
High Efficiency ◽  
Stop Codon ◽  
Signal Sequence ◽  
Foot And Mouth Disease ◽  
High Growth ◽  
High Growth Rate ◽  
Secretory Expression ◽  
Mouth Disease Virus ◽  
Ethanol Yields

Abstract BackgroundHemicelluloses act as one factor contributing to the recalcitrance of lignocelluloses that prevent cellulases to degrade the cellulose efficiently even in low quantities. Supplement of hemicellulases can enhance performance of commercial cellulases in the enzymatic hydrolyses of lignocellulose. Kluyveromyce marxianu is an attractive yeast for cellulosic ethanol fermentation, as well as a promising host for heterologous protein production, since it has remarkable thermotolerance, high growth rate, and broad substrate spectrum etc. In this study, we attempted to coordinately express multiple hemicellulases in K. marxianus through a 2A-mediated ribosomes skipping to self-cleave polyproteins, and investigated their capabilities for saccharification and ethanol production from corncobs.ResultsTwo polycistronic genes IMPX and IMPαX were constructed to test the self-cleavage of P2A sequence from Foot and Mouth Disease virus (FMDV) in K. marxianus. The IMPX gene consisted of a β-mannanase gene M330 (without the stop codon), a P2A sequence and a β-xylanase gene Xyn-CDBFV in turn, while in the IMPαX gene there was an additional α-factor signal sequence in frame with the N-terminus of Xyn-CDBFV. The extracellular β-mannanase activities of IMPX and IMPαX strains were 21.34 and 15.50 U/mL repectively. By contrast, the IMPαX strain secreted 136.17 U/mLof the β-xylanase, which was much higher than that of IMPX strain 42.07 U/mL. Based on these, two recombinant strains, the IXαR and IMPαXPαR, were constructed to coordinately and secretorily express two xylantic enzymes a β-D-xylosidase RuXyn1 and Xyn-CDBFV, or three hemicellulolytic enzymes including M330, Xyn-CDBFV and RuXyn1. In fed-batch fermentations, extracellular activities of β-xylanase and β-xylosidase in the IMPαX strain were 1664.2 and 0.90 U/mL, while productions of secretory β-mannanase, β-xylanase, and β-xylosidase in the IMPαXPαR strain were 159.8, 2210.5, and 1.25 U/ml of respectively. Hemicellulolytic enzymes of these two strains enhanced the yields of both glucose and xylose from diluted acid pretreated (DAP) corncobs when acted synergistically with commercial cellulases. In hybrid saccharification and fermentation (HSF) of DAP corncobs, hemicellulases of the IMPαXPαR strain increased the ethanol yields by 8.7% at 144 h. When using aqueous ammonia pretreated (AAP) corncobs as HSF feedstocks, the IMPαXPαR strain increased both ethanol and xylose yields, which were about 12.7% and 18.2% more than that of the control at 120 h. Our results indicated that coordinately expression of hemicellulolytic enzymes in K. marxianus could promote the saccharification and ethanol production from corncobs.ConclusionsThe FMDV P2A sequence showed high efficiency in self-cleavage of polyproteins in K. marxianus, and could be used for secretory expression of multiple enzymes in present of their own signal sequences. The IMPαXPαR strain that coexpressed three hemicellulolytic enzymes improved the saccharification and ethanol production from corncobs, and could be used as a promising strain for ethanol production from lignocelluloses.

scholarly journals The Ability of Integrin αvβ3 To Function as a Receptor for Foot-and-Mouth Disease Virus Is Not Dependent on the Presence of Complete Subunit Cytoplasmic Domains

2001 ◽  
Vol 75 (1) ◽  
pp. 527-532 ◽  
Author(s):  
Sherry Neff ◽  
Barry Baxt
Keyword(s):  
Viral Replication ◽  
Disease Virus ◽  
High Efficiency ◽  
Cultured Cells ◽  
Foot And Mouth Disease ◽  
Integrin Αvβ3 ◽  
Mouth Disease ◽  
Cytoplasmic Domains ◽  
Mouth Disease Virus ◽  
Foot And Mouth

ABSTRACT The integrin αvβ3 has been shown to function as one of the integrin receptors on cultured cells for foot-and-mouth disease virus (FMDV), and high-efficiency utilization of the bovine homolog of this integrin is dependent on the cysteine-rich repeat region of the bovine β3 subunit. In this study we have examined the role of the cytoplasmic domains of the αv and β3 subunits in FMDV infection. We have found that truncations or extensions of these domains of either subunit, including deletions removing almost all of the cytoplasmic domains, had little or no effect on the ability of the integrin to function as a receptor for FMDV. The lysosomotropic agent monensin inhibited viral replication in cells transfected with either intact or cytoplasmic domain-truncated αvβ3. In addition, viral replication in transfected cells was inhibited by an αvβ3 function-blocking antibody but not by function-blocking antibodies to three other RGD-directed integrins, suggesting that these integrins are not involved in the infectious process. These results indicate that alterations to the cytoplasmic domains of either subunit, which lead to the inability of the integrin receptor to function normally, do not abolish the ability of the integrin to bind and internalize this viral ligand.

scholarly journals effectiveness of indigenous local microorganisms in degrading hexavalent chromium (Cr(VI)) in Batik liquid waste

2021 ◽  
Vol 1 (1) ◽  
pp. 19-29
Author(s):  
Reza Fauzi Dwisandi ◽  
Frista Mutiara ◽  
Elsa Nurfauziah ◽  
Vita Meylani
Keyword(s):  
Hexavalent Chromium ◽  
High Efficiency ◽  
Waste Treatment ◽  
Liquid Waste ◽  
High Growth ◽  
Textile Wastewater ◽  
High Growth Rate ◽  
Gram Negative Bacteria ◽  
Small And Medium Industry

The batik industry in Indonesia has an IKM (Small and Medium Industry) scale so that it does not yet have adequate waste treatment. In the long term, waste is disposed of directly into the environment which can damage aquatic ecosystems and harm human health. Textile wastewater has a complementary picture and has a deep color. One of the most dangerous heavy metals contained in textile waste is hexavalent chromium (Cr(VI)). Several ways can be done to reduce hexavalent chromium (Cr(VI)) by bioremediation. Based on the results of the literature review, it shows that the bioremediation agents from single isolate microorganisms that are most effective in degrading chromium with high efficiency are Bacillus subtilis and Pseudomonas aeruginosa. The most effective consortium servers with constant reduction rates are the consortium of bacteria genus Mesophilobacter, Methylococcus, Agrobacterium, Neisseria, Xanthobacter, Deinococcus, Sporosarcina, and Bacillus by reducing BOD levels by 85.71%. The hexavalent chromium-degrading microorganisms are characterized by the presence of chromate reductase enzymes, mostly gram-negative bacteria, and a high growth rate.

scholarly journals Extension of Flavivirus Protein C Differentially Affects Early RNA Synthesis and Growth in Mammalian and Arthropod Host Cells

2009 ◽  
Vol 83 (21) ◽  
pp. 11201-11210 ◽  
Author(s):  
Sabrina Schrauf ◽  
Christian W. Mandl ◽  
Lesley Bell-Sakyi ◽  
Tim Skern
Keyword(s):  
Host Cell ◽  
Protein C ◽  
Rna Synthesis ◽  
Signal Sequence ◽  
Foot And Mouth Disease ◽  
Rna Replication ◽  
Vero Cells ◽  
Viral Assembly ◽  
Host Cells ◽  
Mouth Disease Virus

ABSTRACT The translation of flaviviral RNA genomes yields a single polyprotein that is processed into the mature proteins by viral and host cell proteases. Mature capsid protein C is freed from the polyprotein by the viral NS2B/3 protease, cleaving in the C-terminal region of protein C in front of the signal sequence for prM. Protein C has been shown to be involved in viral assembly and RNA packaging. To examine further the role of protein C and its production by proteolysis, we replaced the NS2B/3 capsid cleavage site in tick-borne encephalitis virus (TBEV) and West Nile virus (WNV) by the 2A protein of foot-and-mouth disease virus (TBEV-2A and WNV-2A). This obviated the need for NS2B/3 processing at the C terminus of mature protein C while simultaneously producing a 19-amino-acid extension on protein C. Infectious virions were generated with both viruses; the phenotype depended on the host cell. TBEV-2A replicated well in BHK-21 cells but was essentially incapable of replication in tick cells. In contrast, WNV-2A replicated well in mosquito cells but showed a small-plaque phenotype in Vero cells, with frequent production of larger plaques. Sequencing of viral RNA from the larger plaques showed substitutions in the signal sequence for prM, presumably improving coordinated protein processing at the C-prM junction. Furthermore, both TBEV-2A and WNV-2A were also defective in unpackaging and/or early RNA synthesis. Together, these results indicate a role for flavivirus protein C in both viral assembly and RNA replication, possibly by interacting with host cell factors required to set up the cell for RNA replication.

scholarly journals Analysis of a Foot-and-Mouth Disease Virus Type A24 Isolate Containing an SGD Receptor Recognition Site In Vitro and Its Pathogenesis in Cattle

2005 ◽  
Vol 79 (20) ◽  
pp. 12989-12998 ◽  
Author(s):  
Elizabeth Rieder ◽  
Tina Henry ◽  
Hernando Duque ◽  
Barry Baxt
Keyword(s):  
Disease Virus ◽  
High Efficiency ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
Integrin Receptors ◽  
Rgd Sequence ◽  
Mouth Disease Virus ◽  
Receptor Recognition ◽  
Foot And Mouth

ABSTRACT Foot-and-mouth disease virus (FMDV) initiates infection by binding to integrin receptors via an Arg-Gly-Asp (RGD) sequence found in the G-H loop of the structural protein VP1. Following serial passages of a type A24 Cruzeiro virus (A24Cru) in bovine, via tongue inoculation, a virus was generated which contained an SGD sequence in the cell receptor-binding site and expressed a turbid plaque phenotype in BHK-21 cells. Propagation of this virus in these cells resulted in the rapid selection of viruses that grew to higher titers, produced clear plaques, and now contained an RGD sequence in place of the original SGD. To study the role of the SGD sequence in FMDV receptor recognition and bovine virulence, we assembled an infectious cDNA clone of an RGD-containing A24Cru and derived mutant clones containing either SGD with a single nucleotide substitution in the R144 codon or double substitutions at this position to prevent mutation of the S to an R. The SGD viruses grew poorly in BHK-21 cells and stably maintained the sequence during propagation in BHK-21 cells expressing the bovine αVβ6 integrin (BHK3-αVβ6), as well as in experimentally infected and contact steers. While all the SGD-containing viruses used only the bovine αVβ6 integrin as a cellular receptor with relatively high efficiency, the revertant RGD viruses utilized either the αVβ1 or αVβ3 bovine integrins with higher efficiency than αVβ6 and grew well in BHK-21 cells. Replacing the R at the −1 SGD position with either K or E showed that this residue did not contribute to integrin utilization in vitro. These results illustrate the rapid evolution of FMDV with alteration in receptor specificity and suggest that viruses with sequences other than RGD, but closely related to it, can still infect via integrin receptors and induce and transmit the disease to susceptible animals.

Biophysical Measurement of Molecular Weight of Foot-and-Mouth Disease RNA Fragments

1974 ◽  
Vol 32 ◽  
pp. 262-263
Author(s):  
Sydney S. Breese ◽  
Howard L. Bachrach
Keyword(s):  
Chemical Properties ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
Distilled Water ◽  
Bovine Plasma ◽  
Mouth Disease Virus ◽  
Foot And Mouth ◽  
Carbon Coated ◽  
Rna Fragments ◽  
Physical And Chemical

Continuing studies on the physical and chemical properties of foot-and-mouth disease virus (FMDV) have included electron microscopy of RNA strands released when highly purified virus (1) was dialyzed against demlneralized distilled water. The RNA strands were dried on formvar-carbon coated electron microscope screens pretreated with 0.1% bovine plasma albumin in distilled water. At this low salt concentration the RNA strands were extended and were stained with 1% phosphotungstic acid. Random dispersions of strands were recorded on electron micrographs, enlarged to 30,000 or 40,000 X and the lengths measured with a map-measuring wheel. Figure 1 is a typical micrograph and Fig. 2 shows the distributions of strand lengths for the three major types of FMDV (A119 of 6/9/72; C3-Rezende of 1/5/73; and O1-Brugge of 8/24/73.

High resolution surface structure of foot-and-mouth disease virus

1978 ◽  
Vol 36 (2) ◽  
pp. 376-377
Author(s):  
S. S. Breese ◽  
H. L. Bachrach
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Surface Structure ◽  
Disease Virus ◽  
Potassium Phosphate ◽  
Foot And Mouth Disease ◽  
Mouth Disease ◽  
Physical Measurements ◽  
Mouth Disease Virus ◽  
Foot And Mouth

Models for the structure of foot-and-mouth disease virus (FMDV) have been proposed from chemical and physical measurements (Brown, et al., 1970; Talbot and Brown, 1972; Strohmaier and Adam, 1976) and from rotational image-enhancement electron microscopy (Breese, et al., 1965). In this report we examine the surface structure of FMDV particles by high resolution electron microscopy and compare it with that of particles in which the outermost capsid protein VP3 (ca. 30, 000 daltons) has been split into smaller segments, two of which VP3a and VP3b have molecular weights of about 15, 000 daltons (Bachrach, et al., 1975).Highly purified and concentrated type A12, strain 119 FMDV (5 mg/ml) was prepared as previously described (Bachrach, et al., 1964) and stored at 4°C in 0. 2 M KC1-0. 5 M potassium phosphate buffer at pH 7. 5. For electron microscopy, 1. 0 ml samples of purified virus and trypsin-treated virus were dialyzed at 4°C against 0. 2 M NH4OAC at pH 7. 3, deposited onto carbonized formvar-coated copper screens and stained with phosphotungstic acid, pH 7. 3.

The Job Of Microfinance For The Advancement Of Self Help Groups In India

2019 ◽  
Vol 118 (8) ◽  
pp. 236-240
Author(s):  
Dr.R. Murugesan ◽  
M. Leelavathi ◽  
Dr. K. Ravindran
Keyword(s):  
High Growth ◽  
High Growth Rate ◽  
Entrepreneurial Skills ◽  
Self Help ◽  
The Poor ◽  
Total Food ◽  
Self Help Groups ◽  
Positive Step ◽  
Food Consumed

towards jumping from the category of developing economy to developed economy there is one big factor that stops and poses a hindrance in its path of advancement and that obstacle is termed as Poverty. The Indian economic policy focuses on a high growth rate along with a equal participation of the poor so that they avail the opportunities available in the market economy. And in order to ensure the participation of the poor it has become important for the country to create a platform where the poor can easily access the various financial products. Microfinance is one such strategy for inclusive growth. Microfinance can change the life of the poor though not completely but a reasonable change can be ensured. In different phases of life women play a crucial role despite the discrimination that is faced by them. But equality can be endowed to women by enhancing the entrepreneurial skills in them. This is possible through Self Help Groups (SHGs). In India women produce around 30% of the total food consumed but she gets only 10% of the property or wealth of the country. Development of women is inevitable for the development and growth of any economy. SHGs happen to be a positive step in this direction. Along with these mediums there should be a cheap and easy source of credit for them and Microfinance fulfills the requirement. This study aims to find the role of this strong medium of Microfinance in the advancement of SHGs in India

Sign in / Sign up

Export Citation Format

Share Document