Penicillium citrinum UFV1 β-glucosidases: purification, characterization, and application for biomass saccharification

Abstract

Backgroundβ-Glucosidases are components of the cellulase system, a family of enzymes that hydrolyze the β-1,4 linkages of cellulose. These proteins have been extensively studied due to the possibility of their use in various biotechnological processes. They have different affinities for substrates (depending on their source) and their activities can be used for saccharification of different types of biomass. In this context, the properties and the synergistic capacity of β-glucosidases from different organisms, to supplement the available commercial cellulase cocktails, need a comprehensive evaluation.ResultsTwo β-glucosidases belonging to GH3 family were secreted by Penicillium citrinum UFV. PcβGlu1 (241 kDa) and PcβGlu2 (95 kDa) presented acidic and thermo-tolerant characteristics. PcβGlu1 showed Michaelis–Menten kinetics for all substrates tested with Km values ranging from 0.09 ± 0.01 (laminarin) to 1.7 ± 0.1 mM (cellobiose, C2) and kcat values ranging from 0.143 ± 0.005 (laminarin) to 8.0 ± 0.2 s−1 (laminaribiose, Lb). PcβGlu2 showed substrate inhibition for 4-methylumbelliferyl-β-d-glucopyranoside (MUβGlu), p-nitrophenyl-β-d-glucopyranoside (pNPβGlu), cellodextrins (C3, C4, and C5), N-octil-β-d-glucopyranoside, and laminaribiose, with Km values ranging from 0.014 ± 0.001 (MUβGlu) to 0.64 ± 0.06 mM (C2) and kcat values ranging from 0.49 ± 0.01 (gentiobiose) to 1.5 ± 0.2 s−1 (C4). Inhibition constants (Ki) for PcβGlu2 substrate inhibition ranged from 0.69 ± 0.07 (MUβGlu) to 10 ± 1 mM (Lb). Glucose and cellobiose are competitive inhibitors of PcβGlu1 and PcβGlu2 when pNPβGlu is used as a substrate. For PcβGlu1 inhibition, Ki = 1.89 ± 0.08 mM (glucose) and Ki = 3.8 ± 0.1 mM (cellobiose); for PcβGlu2, Ki = 0.83 ± 0.05 mM (glucose) and Ki = 0.95 ± 0.07 mM (cellobiose). The enzymes were tested for saccharification of different biomasses, individually or supplementing a Trichoderma reesei commercial cellulose preparation. PcβGlu2 was able to hydrolyze banana pseudostem and coconut fiber with the same efficiency as the T. reesei cocktail, showing significant synergistic properties with T. reesei enzymes in the hydrolysis of these alternative biomasses.ConclusionsThe β-glucosidases from P. citrinum UFV1 present different enzymatic properties from each other and might have potential application in several biotechnological processes, such as hydrolysis of different types of biomass.Electronic supplementary materialThe online version of this article (10.1186/s13068-018-1226-5) contains supplementary material, which is available to authorized users.

Cellulase Addition and Pre-hydrolysis Effect of High Solid Fed-Batch Simultaneous Saccharification and Ethanol Fermentation from a Combined Pretreated Oil Palm Trunk

“…Endoglucanase exposes the polymer to cellobiohydrolase by randomly attacking the amorphous sites of cellulose . Cellobiose and short oligosaccharides inhibit the activity of endoglucanase and cellobiohydrolase, whereas β-glucosidase is accountable for the rate-limiting step of the entire cellulolytic process and glucose production . Until 36 h of PSSF, the ethanol quantity increased; conversely, it later decreased owing to enough substrate presence for exponential growth of SC90.…”

Isolation and Molecular Characterization of Indigenous Penicillium chrysogenum/rubens Strain Portfolio for Penicillin V Production

“…Moreover, P. chrysogenum clade produces secalonic acids D, F, and lumpidin-like compounds along with metabolites produced by P. rubens strains [ 4 ]. Penicillium strains are known for producing essential molecules, including enzymes, organic acids, terpenoids, and polyketides used in various industries ( Table 3 ) [ 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 , 51 , 52 , 53 , 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 , 62 , 63 , 64 , 65 , 66 , 67 , 68 , 69 , 70 ].…”

Fungal Dysbiosis Correlates With the Development of Tumour-Induced Cachexia in Mice

“…Finally, it is worth mentioning that the population analysis shown in Figure 4 allowed identification of four fungal species preferably present in the gut of SC animals, which can be grown under axenic laboratory conditions (Rhizopus oryzae, Cystobasidium sloofiae, Penicillium citrinum and Cladiosporium halotolerans). These fungi are natural candidates to be tested (individually, or in combinations) for their eventual roles as probiotic agents, aiming at the prevention/treatment of cachexia (76,108). Rhyzopus oryzae, in particular, stands out as the most promising probiotic candidate, not only due to its capacity to produce bioactive substances, such as chitosan and gallic acid, but also to its historical use as a component of traditional oriental foods, which has granted it the status of a GRAS (Generally Recognized As Safe) filamentous fungus, according to the US Food and Drug Administration (FDA) (76).…”