Cellulosomal hemicellulases:Indispensable players for ensuring effective lignocellulose bioconversion

The bioconversion of lignocellulose has attracted global attention,due to the significant potential of agricultural and forestry wastes as renewable zero-carbon resources and the urgent need for substituting fossil carbon.The cellulosome system is a multi-enzyme complex produced by anaerobic bacteria,which comprises cellulases,hemicellulases,and associated enzymatic and non-enzymatic components that promote biomass conversion.To enhance their efficiency in degrading recalcitrant lignocellulosic matrices,cellulosomes have been employed to construct biocatalysts for lignocellulose bioconversion,such as consolidated bioprocessing and consolidated bio-saccharification.Hemicelluloses,the second most abundant polysaccharides in plant cell walls,hold valuable application potential but can also induce inhibitory effects on cellulose hydrolysis,thus highlighting the indispensable roles of hemicellulases within the cellulosome complex.This review evaluated current research on cellulosomal hemicellulases,comparing their types,abundance,and regulation,primarily focusing on eight known cellulosome-producing species of different origins.We also reviewed their growth conditions,their hemicellulose-degrading capabilities,and the inhibitory effects of hemicellulose on cellulosome-based lignocellulose saccharification.Finally,we proposed strategies for targeted enhancement of hemicellulase in cellulosomes to improve lignocellulose bioconversion in future studies.

Isolation of protoplast from callus of Populus euphratica and H^+ fluxes across plasma membrane under NaCl stress

We used callus of Populus euphratica Olive to isolate protoplasts, and IT fluxes across plasma membrane were investigated. The concentration of enzymes for protoplast isolation, e.g. cellulase, pectolyase, macerozyme, hemicellulase, and sorbitol content, incubation time were systemically studied. High yield and viability of protoplast was achieved after 6-8 hours incubation of P. euphratica callus in enzyme solution containing 1.5% （w：v） cellulase R-10, 0.1% （w：v） pectolyase Y-23, 0.2% （w：v） macerozyme R-10, 0.05% （w：v） hemicellulase and 0.75M）.80 mol·L^-1 sorbitol. Non-invasively ion selective microelectrode technique was used to access proton fluxes in the absence and presence of NaCl （20 mmol.L-1）. Salt-induced transient net IT effiux was observed in the plasma membrane ofP. euphratica cells. The shift of IT flux response to NaC1 shock and the relevance to salt tolerance were discussed.

A comparative analysis of β-mannanases of bacteria from Antarctica and Malaysia

β-mannanase is an enzyme that is commonly expressed in environmental bacteria. It degrades hemicellulose found in plant material and recycles nutrients back into the environment. Because this enzyme significantly contributes to biodegradation and has recently been applied in industry, we conducted a comparative analysis of bacterial isolates found in soil samples from Schirmacher Oasis, Antarctica, and Sabah, Malaysia that were capable of degrading mannan. A total of 9 bacterial isolates from Antarctica and 30 bacterial isolates from Malaysia exhibited β-mannanase activity. These bacteria were differentiated and clustered using their random amplified polymorphic DNA （RAPD） profiles, and the β-mannanase activity of these isolates was tested at different temperatures and pH. Five out of 9 Antarctica isolates and seven out of 30 Malaysian isolates were identified based on their 16S rDNA sequences. Identified bacterial isolates from Antarctica were： MP1 （Bacillus amyloliquefaciens）, MP2 （Bacillus pumilus）, MP5 （Bacilluspumilus）, A40 （Arthrobacter sp.）, and C27 （Arthrobacter oxydans）. Identified bacterial isolates from Ma- laysia were： Y1 （Paenibacillus sp.）, Y2 （Bacillus sp.）, Y16 （Paenibacillus sp.）, Y18 （Paenibacillus sp.）, A7 （Paenibacillus sp.）, B26 （Streptomyces sp.）, and D4 （Paenibacillus amylolyticus）. β-mannanases produced by the Antarctica bacterial isolates MP1 （Bacillus amyloliquefaciens） and A40 （Arthrobacter sp.） were active at 5℃ and 20℃, respectively, while the β-mannanase pro- duced by the bacterial isolate from Malaysia, A7 （Paenibacillus sp.）, was active at 35 ℃.

Integrated engineering of enzymes and microorganisms for improving the efficiency of industrial lignocellulose deconstruction

Bioconversion of lignocellulosic biomass to fuels and chemicals represents a new manufacturing paradigm that can help address society’s energy,resource,and environmental problems.However,the low efficiency and high cost of lignocellulolytic enzymes currently used hinder their use in the industrial deconstruction of lignocellu-lose.To overcome these challenges,research efforts have focused on engineering the properties,synergy,and production of lignocellulolytic enzymes.First,lignocellulolytic enzymes’catalytic efficiency,stability,and toler-ance to inhibitory compounds have been improved through enzyme mining and engineering.Second,synergistic actions between different enzyme components have been strengthened to construct customized enzyme cocktails for the degradation of specific lignocellulosic substrates.Third,biological processes for protein synthesis and cell morphogenesis in microorganisms have been engineered to achieve a high level and low-cost production of lignocellulolytic enzymes.In this review,the relevant progresses and challenges in these fields are summa-rized.Integrated engineering is proposed to be essential to achieve cost-effective enzymatic deconstruction of lignocellulose in the future.

Monitoring the fibrolytic potential of microbial ecosystems from domestic and wild ruminants browsing tanniferous forages

Although the rumen microbiome has been reported to synthesize a rich source of symbiotic enzymes(exocellulase, endocellulase, hemicellulase and cellobiase), the digestion of tropical C_4 grasses and browses by ruminants is still limited. Therefore, this study aimed to unveil potential fibrolytic microbial ecosystems from giraffe, kudu, impala and consortia(A1 [giraffe + kudu], A2 [giraffe + impala], A3[kudu + impala], and A4 [giraffe + kudu + impala]) browsing tanniferous plants, which can be used to improve forage utilization in domesticated goat. Crude protein enzyme extracts(CPZ) from fresh faecal samples were precipitated by 60% ammonium sulphate and assayed for exocellulase, endocellulase and hemicellulase by incubating with crystalline cellulose, carboxymethyl cellulose and xylan at 38 ℃ with optimum pH of 5.5 to 6.5 for 1, 2, and 48 h, respectively. Enzyme specific activities were defined as μg of reducing sugar/mg CPZ. In vitro fermentation study was done by transferring 33 mL of fresh faecal inoculum into 67 mL of salivary buffer containing 1 g Acacia sieberiana and incubating for 72 h at 38 ℃,Apparent degradability(APDeg), true degradability(TD), neutral detergent fibre degradability(NDFdeg),acid detergent fibre degradability(ADFdeg), microbial yield(MY), metabolizable energy(ME) and total gas emitted(Gas) were measured. Exocellulase activities were higher(P < 0.05) in all wild animals and consortia than those in goat except for A4. Minimal differences in hemicellulase activities(P < 0.05) were observed among goat and wild animals and consortia, while endocellulase activity was generally higher(P < 0.05) in goat than that in the rest of the systems. Apart from A3, TDeg, NDFdeg and ADFdeg were higher(P < 0.05) in all microbial ecosystems from wild animals and consortia than those in goat.Apparent degradability, MY and ME also varied(P < 0.05) among these systems. Giraffe, Kudu and A3 produced lower(P < 0.05) gas than the goat system. This study showed that microbial ecosystems from wild browsers(especially impala) and consortia possess a higher potential to digest tanniferous forage with less enteric gas production compared with domesticated goat, hence those microbiome could be exploited as microbial feed additives for improving digestibility and reducing enteric gas production in domesticated goat. Improvements of goat's digestibility will depend on the survival and establishment of microbial species in the rumen as well as their fibrolytic and symbiotic potential including tannin tolerance.