scholarly journals Cellulose hydrolysis by the cellulases from Trichoderma reesei: a new model for synergistic interaction

1994 ◽  
Vol 298 (3) ◽  
pp. 705-710 ◽  
Author(s):  
B Nidetzky ◽  
W Steiner ◽  
M Hayn ◽  
M Claeyssens
Keyword(s):  
Trichoderma Reesei ◽  
Filter Paper ◽  
Cellulose Hydrolysis ◽  
Catalytic Core ◽  
Core Proteins ◽  
Hydrolysis Rates ◽  
Cellulose Binding ◽  
The Mathematical Model ◽  
Hydrolysis Of ◽  
Synergistic Combinations

The hydrolysis of Whatman no. 1 filter paper by purified cellulolytic components from Trichoderma reesei and the synergistic action of binary combinations of these enzymes on the same substrate were investigated. At 20 milligrams filter paper, enzyme concentrations needed to obtain half-maximal hydrolysis rates (KE values) were in the 3-4 microM range for the cellobiohydrolases (CBHs) and 0.05-0.10 microM for the endoglucanases (EGs). Catalytic-core proteins of CBH I and EG III, lacking the cellulose-binding domain, exhibit KE values 2.3 and 5.1 times higher than those of the intact enzymes. In synergistic combinations of two cellulases, the KE value of at least one enzyme was 3-10-fold reduced. CBH I/CBH II and CBH I/EG III combinations showed the most powerful synergism, and optimal ratios were a function of the total protein concentration. Results obtained in activity and adsorption assays using filter paper pretreated with one component, followed by inactivation and subsequent hydrolysis with the same or another cellulase component, point to a sequential enzymic attack of the cellulose and seems consistent with the mathematical model presented.

scholarly journals Cellulose hydrolysis by the cellulases from Trichoderma reesei: adsorptions of two cellobiohydrolases, two endocellulases and their core proteins on filter paper and their relation to hydrolysis

1994 ◽  
Vol 303 (3) ◽  
pp. 817-823 ◽  
Author(s):  
B Nidetzky ◽  
W Steiner ◽  
M Claeyssens
Keyword(s):  
Mass Transfer ◽  
Trichoderma Reesei ◽  
Filter Paper ◽  
Cellulose Hydrolysis ◽  
Solid Substrate ◽  
Bulk Liquid ◽  
Cellobiohydrolase I ◽  
Irreversible Adsorption ◽  
Core Proteins ◽  
Cellulose Binding

Separate binding of several purified cellulolytic components of Trichoderma reesei on to filter paper was studied and concomitant hydrolysis rates evaluated. Enhancement of mass transfer from the bulk liquid to the solid substrate by agitation has two different effects on adsorption depending on the type of enzyme: (i) the fraction of cellobiohydrolase II (CBH II) and endoglucanase III (EG III) bound at equilibrium is increased, whereas (ii) the rate but not the extent of cellobiohydrolase I (CBH I) and endoglucanase I (EG I) adsorption is affected. The adsorption of CBH I core, a component lacking the cellulose-binding domain (CBD), is, however, not significantly influenced by mass transfer. The CBH I interdomain peptide (present in CBH I core b) does not participate in adsorption but enhances stability. The adsorption of CBH I core proteins is a fully reversible process whereas that of the intact CBH I is not. Thus, the interaction of the CBD with filter paper apparently accounts for the mass-transfer-limited binding rate and also for the irreversible adsorption of intact CBH I. Adsorption isotherms at 50 degrees C indicate very similar relative association constants for the intact cellulases (0.24-0.30 l/g of cellulose), but drastically reduced values for CBH I core proteins (0.03 l/g of cellulose). The specific activities of adsorbed CBH I and of its core proteins are identical and a linear relationship between adsorption and rates of hydrolysis is found only for these enzymes. Thus, non-productive binding on to cellulose seems evident in the case of CBH II and EG III but not CBH I.

A cellulose-binding module of the Trichoderma reesei β-mannanase Man5A increases the mannan-hydrolysis of complex substrates

2003 ◽  
Vol 101 (1) ◽  
pp. 37-48 ◽  
Author(s):  
Per Hägglund ◽  
Torny Eriksson ◽  
Anna Collén ◽  
Wim Nerinckx ◽  
Marc Claeyssens ◽  
...  
Keyword(s):  
Trichoderma Reesei ◽  
Cellulose Binding Module ◽  
Cellulose Binding ◽  
Hydrolysis Of

scholarly journals Roles of the Catalytic Domain and Two Cellulose Binding Domains of Thermomonospora fusca E4 in Cellulose Hydrolysis

1998 ◽  
Vol 180 (7) ◽  
pp. 1709-1714 ◽  
Author(s):  
Diana Irwin ◽  
Dong-Hoon Shin ◽  
Sheng Zhang ◽  
Brian K. Barr ◽  
Joshua Sakon ◽  
...  
Keyword(s):  
Filter Paper ◽  
Recombinant Dna ◽  
Catalytic Domain ◽  
Cellulose Hydrolysis ◽  
Hydrolytic Activity ◽  
Thermomonospora Fusca ◽  
Binding Domains ◽  
The Family ◽  
Recombinant Dna Techniques ◽  
Cellulose Binding

ABSTRACT Thermomonospora fusca E4 is an unusual 90.4-kDa endocellulase comprised of a catalytic domain (CD), an internal family IIIc cellulose binding domain (CBD), a fibronectinlike domain, and a family II CBD. Constructs containing the CD alone (E4-51), the CD plus the family IIIc CBD (E4-68), and the CD plus the fibronectinlike domain plus the family II CBD (E4-74) were made by using recombinant DNA techniques. The activities of each purified protein on bacterial microcrystalline cellulose (BMCC), filter paper, swollen cellulose, and carboxymethyl cellulose were measured. Only the whole enzyme, E4-90, could reach the target digestion of 4.5% on filter paper. Removal of the internal family IIIc CBD (E4-51 and E4-74) decreased activity markedly on every substrate. E4-74 did bind to BMCC but had almost no hydrolytic activity, while E4-68 retained 32% of the activity on BMCC even though it did not bind. A low-activity mutant of one of the catalytic bases, E4-68 (Asp55Cys), did bind to BMCC, although E4-51 (Asp55Cys) did not. The ratios of soluble to insoluble reducing sugar produced after filter paper hydrolysis by E4-90, E4-68, E4-74, and E4-51 were 6.9, 3.5, 1.3, and 0.6, respectively, indicating that the family IIIc CBD is important for E4 processivity.

scholarly journals Dynamic Interaction of Trichoderma reesei Cellobiohydrolases Cel6A and Cel7A and Cellulose at Equilibrium and during Hydrolysis

1999 ◽  
Vol 65 (12) ◽  
pp. 5229-5233 ◽  
Author(s):  
Hetti Palonen ◽  
Maija Tenkanen ◽  
Markus Linder
Keyword(s):  
Trichoderma Reesei ◽  
High Performance ◽  
Initial Step ◽  
Cellulose Hydrolysis ◽  
Reductive Methylation ◽  
Intact Proteins ◽  
Catalytic Domains ◽  
Binding Domains ◽  
Binding Isotherms ◽  
Cellulose Binding

ABSTRACT The binding of cellobiohydrolases to cellulose is a crucial initial step in cellulose hydrolysis. In the search for a detailed understanding of the function of cellobiohydrolases, much information concerning how the enzymes and their constituent catalytic and cellulose-binding domains interact with cellulose and with each other and how binding changes during hydrolysis is still needed. In this study we used tritium labeling by reductive methylation to monitor binding of the two Trichoderma reesei cellobiohydrolases, Cel6A and Cel7A (formerly CBHII and CBHI), and their catalytic domains. Measuring hydrolysis by high-performance liquid chromatography and measuring binding by scintillation counting allowed us to correlate activity and binding as a function of the extent of degradation. These experiments showed that the density of bound protein increased with both Cel6A and Cel7A as hydrolysis proceeded, in such a way that the adsorption points moved off the initial binding isotherms. We also compared the affinities of the cellulose-binding domains and the catalytic domains to the affinities of the intact proteins and found that in each case the affinity of the enzyme was determined by the linkage between the catalytic and cellulose-binding domains. Desorption of Cel6A by dilution of the sample showed hysteresis (60 to 70% reversible); in contrast, desorption of Cel7A did not show hysteresis and was more than 90% reversible. These findings showed that the two enzymes differ with respect to the reversibility of binding.

DISTRIBUTION OF HEXOSAMINE IN ELEGTROPHORETIC FRACTIONS OF AVIAN, BOVINE, PORCINE, AND HUMAN SERA

1967 ◽  
Vol 45 (4) ◽  
pp. 541-550 ◽  
Author(s):  
A. P. Gaunce ◽  
P. A. Anastassiadis
Keyword(s):  
Human Blood ◽  
Filter Paper ◽  
Cationic Resin ◽  
Human Sera ◽  
Hydrolysis Of ◽  
Direct Hydrolysis

The distribution of hexosamine among the proteins of avian, bovine, porcine, and human blood sera was studied by electrophoresis on filter paper. Hexosamine was determined after direct hydrolysis of stained sections of the paper, followed by chromatography of hydrolysates on cationic resin. Some substantial and statistically significant differences in hexosamine and protein contents of the zones were found among species.

Effect of Rhodococcus sp. pretreatment on cellulose hydrolysis of corn stalk

2020 ◽  
pp. 1-7
Author(s):  
Pan Hu ◽  
Huanan Li ◽  
Wenjing Xiao ◽  
Xiaohang Xie ◽  
Yuxian Yang ◽  
...  
Keyword(s):  
Cellulose Hydrolysis ◽  
Corn Stalk ◽  
Hydrolysis Of ◽  
Rhodococcus Sp

scholarly journals Improvement of Enzymatic Saccharification of Cellulose-Containing Raw Materials Using Aspergillus niger

Processes ◽  
2021 ◽  
Vol 9 (8) ◽  
pp. 1360
Author(s):  
Ekaterina Budenkova ◽  
Stanislav Sukhikh ◽  
Svetlana Ivanova ◽  
Olga Babich ◽  
Vyacheslav Dolganyuk ◽  
...  
Keyword(s):  
Enzymatic Hydrolysis ◽  
Aspergillus Niger ◽  
Filter Paper ◽  
Raw Materials ◽  
Enzymatic Saccharification ◽  
Wood Chip ◽  
Cellulase Activity ◽  
Wood Chips ◽  
Acid Pretreatment ◽  
Hydrolysis Of

Enzymatic hydrolysis of cellulose-containing raw materials, using Aspergillus niger, were studied. Filter paper, secondary cellulose-containing or starch-containing raw materials, miscanthus cellulose after alkaline or acid pretreatment, and wood chip cellulose, were used as substrates. The study focused on a wild A. niger strain, treated, or not (control), by ultraviolet (UV) irradiations for 45, 60, or 120 min (UV45, UV60, or UV120), or by UV irradiation for 120 min followed by a chemical treatment with NaN3 + ItBr for 30 min or 80 min (UV120 + CH30 or UV120 + CH80). A mixture of all the A. niger strains (MIX) was also tested. A citrate buffer, at 50 mM, wasthe most suitable for enzymatic hydrolysis. As the UV exposure time increased to 2 h, the cellulase activity of the surviving culturewas increased (r = 0.706; p < 0.05). The enzymatic activities of the obtained strains, towards miscanthus cellulose, wood chips, and filter paper, were inferior to those obtained with commercial enzymes (8.6 versus 9.1 IU), in some cases. Under stationary hydrolysis at 37 °C, pH = 4.7, the enzymatic activity of A. niger UV120 + CH30 was 24.9 IU. The enzymatic hydrolysis of secondary raw materials, using treated A. niger strains, was themost effective at 37 °C. Similarly, the most effective treatment of miscanthus cellulose and wood chips occurred at 50 °C. The maximum conversion of cellulose to glucose was observed using miscanthus cellulose (with alkaline pretreatment), and the minimum conversion was observed when using wood chips. The greatest value of cellulase activity was evidenced in the starch-containing raw materials, indicating that A. niger can ferment not only through cellulase activity, but also via an amylolytic one.

scholarly journals Processive action of cellobiohydrolase Cel7A from Trichoderma reesei is revealed as ‘burst’ kinetics on fluorescent polymeric model substrates

2005 ◽  
Vol 385 (2) ◽  
pp. 527-535 ◽  
Author(s):  
Kalle KIPPER ◽  
Priit VÄLJAMÄE ◽  
Gunnar JOHANSSON
Keyword(s):  
Bacterial Cellulose ◽  
Trichoderma Reesei ◽  
Anthranilic Acid ◽  
Degree Of Polymerization ◽  
Cellulose Substrates ◽  
Quantitative Monitoring ◽  
End Groups ◽  
Alternative Means ◽  
Burst Kinetics ◽  
Hydrolysis Of

Reaction conditions for the reducing-end-specific derivatization of cellulose substrates with the fluorogenic compound, anthranilic acid, have been established. Hydrolysis of fluorescence-labelled celluloses by cellobiohydrolase Cel7A from Trichoderma reesei was consistent with the active-site titration kinetics (burst kinetics), which allowed the quantification of the processivity of the enzyme. The processivity values of 88±10, 42±10 and 34±2.0 cellobiose units were found for Cel7A acting on labelled bacterial cellulose, bacterial microcrystalline cellulose and endoglucanase-pretreated bacterial cellulose respectively. The anthranilic acid derivatization also provides an alternative means for estimating the average degree of polymerization of cellulose and, furthermore, allows the quantitative monitoring of the production of reducing end groups on solid cellulose on hydrolysis by cellulases. Hydrolysis of bacterial cellulose by cellulases from T. reesei revealed that, by contrast with endoglucanase Cel5A, neither cellobiohydrolases Cel7A nor Cel6A produced detectable amounts of new reducing end groups on residual cellulose.

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