福寿螺和田螺消化酶活性比较

为比较福寿螺(Pomacea canaliculata(Lamarck,1828))和当地中国圆田螺(Cipangopaludina chinensis(Gray,1832))消化能力的差异,探索福寿螺成功入侵的机制,以田螺为对照,测定了1—4龄的福寿螺和田螺的胃和肝脏的消化酶——纤维素酶(羧甲基纤维素法)、淀粉酶(3,5-二硝基水杨酸法)和脂肪酶(滴定法)的活性。结果表明:1)相同年龄的福寿螺胃和肝脏中的消化酶活性明显高于田螺。其中,纤维素酶活性分别高出1.00—2.11倍、1.66—2.84倍;淀粉酶活性分别高出1.53—3.47倍、1.47—1.80倍;脂肪酶活性分别高出2.07—4.73倍、6.13—9.93倍。2)在生长发育过程中,福寿螺胃和肝脏中的消化酶活性变化幅度(51.2%—131.2%)明显高于田螺(23.3%—47.1%)。3)福寿螺的各种消化酶之间存在协同作用。如福寿螺的淀粉酶活性与脂肪酶活性呈极显著正相关(胃中r=0.736**、肝脏中r=0.867**)。此外,胃中的淀粉酶活性还与纤维素酶活性呈显著正相关关系(r=0.696*)。相应地,田螺胃中的淀粉酶和脂肪酶之间也存在显著的正相关关系(r=0.706*),而肝脏中的纤维素酶与脂肪酶活性呈显著负相关(r=-0.593*)。4)福寿螺对纤维素类和淀粉类物质都有较强的消化能力,且能较好地消化脂肪类物质,而田螺能消化纤维素类和淀粉类物质,对脂肪的消化能力却很弱。福寿螺的纤维素酶和淀粉酶活性分别是田螺的2.42和1.88倍,脂肪酶活性达到了5.66倍。可见,福寿螺具有较高的消化酶活性,且各消化酶之间存在正协同性。这可能是导致福寿螺食量大、食性杂,使其能快速生长和成功入侵的重要原因之一。     

The role of fungi in the diet of the amphipod Gammarus pulex (L.): an enzymatic study

SUMMARY. 1. Sets of ten Gammarus pulex fed on controlled diets of sterile alder leaves, or fungal mycelium, or alder leaves incubated for 10 days with an aquatic hyphomycete, were assayed for cellulase, β-1,3-glucanase an d chiitinase activity and compared with (a) animals taken directly from the stream, (b) animals starved for 2 days, and (c) enzyme activity in fungal mycelium.
2. Gut enzyme activity was compared on natural substrates of sterile leaves, mycelium and inoculated leaves as well as on model substrates.
3. G. pulex secretes an endogenous coupled cellulase system capable of degrading native cellulose in plant cell walls. It also secretes β-1,3-glucanase and chitinase capable of degrading fungal cell walls thus affording access for gut enzymes to cell contents.
4. Secretion of enzymes active on native cellulose is enhanced on a diet of leaves already partially degraded by fungal enzymes. Gut enzymes extract more reducing sugar from this substrate than from sterile leaves. Specific enzyme secretion is enhanced by the presence in the diet of exposed, accessible substrates. Fungal enzymes do not appear to contribute to the digestive processes of G. pulex.     

Digestive enzymes of the saltmarsh periwinkleLittorina irrorata (Mollusca: Gastropoda)

Summary The saltmarsh periwinkleLittorina irrorata is well adapted for the digestion of a wide range of polysaccharides. Enzyme extracts attacked cellulose, pectin, xylan, bean gum and mannan (common in cell walls of higher plants), as well as starch and laminarin (representative of major polysaccharide classes in fungal, algal, and animal tissues). Activities were generally highes at a ph of 5 or 6. There was no evidence that chitin was digested, but 19 other enzymes, active toward various carbohydrates, lipids and peptides, were demonstrated. Enzymatic activity toward Azocoll, a general substrate for proteinase activity, was weak compared to that of other aquatic detritivores. The maximum was reached at a pH of 8. Enzymatic activities were generally measured with extracts of the entire visceral hump. Separate stomach or intestine extracts also gave strong activities. The stomach was the most acidic section of the digestive system with an average pH of 5.8; the intestine had an average pH of 7.3.     

Rapid isolation and high-throughput determination of cellulase and laminarinase activity in soils

A new method for extracting soil enzymes is described and a microplate method for assaying soil β-1,4-glucanases (cellulases) and β-1,3-glucanases (laminarinases). Soil samples were mechanically disrupted to produce crude enzyme extracts, and diluted preps incubated in microplates containing either carboxymethyl cellulose (CMC) to determine cellulase activity or laminarin substrate to determine laminarinase activity. The resulting glucose was measured using the fluorometric Amplex Red^® glucose assay. The method was reproducible, could be completed in 1 day and measured twice as much enzyme activity than the standard passive soil enzyme extraction procedure. The method described herein facilitates the development of high-throughput soil multiplex enzymatic assays from several soil samples at one time, and is well suited to the study of functional microbial ecology.     

Purification and partial characterization of a novel hyaluronic acid-degrading enzyme from Antarctic krill (Euphausia superba)

Summary A novel enzyme degrading hyaluronic acid has been isolated, purified and characterized from Antarctic krill (Euphausia superba). A combination of affinity chromatography (Con A-Sepharose), gel filtration (Superose 6) and fast protein liquid chromatography (Mono Q) was used for the purification. The hyaluronidase activity was determined by a radial diffusion method based on hyaluronic acid incorporated into an agarose gel. Moreover, the beta-glucuronidase and endo-(1,3)-beta-D-glucanase activities were also followed through the process using phenolphtalein mono beta-glucuronic acid and laminarin as substrates. After the final purification step on Mono Q column, the chromatogram showed three main peaks designated A, B and C. Peak C contained high hyaluronidase activity undetectable in peak A and B. The betaglucuronidase activity was associated with peak A, while the endo-(1,3)-beta-D-glucanase activity was found in peak B and slight in peak C. The hyaluronidase was purified about 85-fold. It had a pH optimum of 5.3, a temperature optimum of 37°C and a molecular weight of 80 000 Daltons. On polyacrylamide gradient gel electrophoresis the enzyme fraction showed one major band associated with hyaluronic acid decomposition, slightly contaminated with a few other components. Isoelectric focusing in combination with a hyaluronic acid zymogram demonstrated one major band at pH 6.7 with high enzyme activity. Preliminary data on enzyme specificity suggest that krill hyaluronidase is a new endo-beta-glucuronidase and support the concept of krill enzymes as a remarkable and unusually effective digestive system adapted to the Antarctic marine ecosystem.     

Dietary preference and digestive enzyme activities as indicators of trophic resource utilization by six species of crab

The digestive physiology and stomach contents of six crab species from a variety of habitats were investigated to provide an indication of their digestive capability and dietary preferences. Stomach contents varied between species, but the key enzymes present were generally consistent with the types of dietary material being ingested. Nectocarcinus integrifons (red rock crab) consumed large quantities of seagrass and had high cellulase activity (0.02+/-0.004 units mg-1) to digest the constituent cellulose. Petrolisthes elongatus (porcelain crab) ingested brown and green phytoplankton and algae and had considerable laminarinase (0.35+/-0.08 units mg-1) and beta-glucosidase (0.025+/-0.005 units mg-1) activities to digest the laminarin in its diet. Leptograpsus variegatus (omnivorous swift-footed shore crab) had high activities of protease (1.2+/-0.02 units mg-1), alpha-glucosidase, and alpha-amylase and appeared well equipped to utilize both dietary protein and carbohydrate. Stomach contents in Nectocarcinus tuberculosus (velvet crab) and Carcinus maenas (green crab) also suggest that these species are omnivorous. N. tuberculosus had high cellulase and chitinase for digesting the cellulose in plants and the chitin in invertebrate shells respectively. C. maenas had intermediate digestive enzyme levels and may employ more of a generalist feeding strategy than other species. Plagusia chabrus (speedy crab) is carnivorous, consuming encrusting bryozoans, hydroids, crustaceans, and fish. It has high protease activity, particularly trypsin (0.73+/-0.12 units mg-1), to digest the protein in its animal prey. Each species of crab studied had a complex suite of digestive enzymes, the relative activities of which reflected individual and very different species-specific dietary niches.     

Enzymatic degradation of steam-pretreated Lespedeza stalk (Lespedeza crytobotrya) by cellulosic-substrate induced cellulases

The cellulase production by Trichoderma viride, cultivated on different substrates, namely steam-pretreated Lespedeza, filter paper, microcrystalline cellulose (MCC) or carboxymethyl cellulose (CMC), was studied. Different cellulase systems were secreted when cultivated on different substrates. The cellulolytic enzyme from steam-pretreated Lespedeza medium performed the highest filter paper activity, exoglucanase and endoglucanase activities, while the highest β-glucosidase activity was obtained from the enzyme produced on filter paper medium. The hydrolytic potential of the enzymes produced from different media was evaluated on steam-pretreated Lespedeza. The cellulase from steam-pretreated Lespedeza was found to have the most efficient hydrolysis capability to this specific substrate. The molecular weights of the cellulases produced on steam-pretreated Lespedeza, filter paper and MCC media were 33, 37 and 40 kDa, respectively, and the cellulase from CMC medium had molecular weights of 20 and 43 kDa. The degree of polymerization, crystallinity index and micro structure scanned by the scanning electron microscopy of degraded steam-pretreated Lespedeza residues were also studied.     

Isolation and properties of a (1,3)-beta-D-glucanase from Ruminococcus flavefaciens

A (1,3)-beta-D-glucanase [(1,3)-beta-D-glucan-3-glucanohydrolase] from Ruminococcus flavefaciens grown on milled filter paper was purified 3,700-fold (19% yield) and appeared as a single major protein and activity band upon polyacrylamide gel electrophoresis. The enzyme did not hydrolyze 1,6-beta linkages (pustulan) or 1,3-beta linkages in glucans with frequent 1,6-beta-linkage branch points (scleroglucan). Curdlan and carboxymethylpachyman were hydrolyzed at 50% the rate of laminarin. The enzyme had a Km of 0.37 mg of laminarin per ml, a pH optimum of 6.8, and a temperature optimum of 55 degrees C and was stable to heating at 40 degrees C for 60 min. The molecular mass of the enzyme was estimated to be 26 kDa by gel filtration and 25 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme was completely inhibited by 1 mM Hg2+, Cu2+, and KMnO4, 75% by 1 mM Ag2+, and Ni2+, and 50% by 1 mM Mn2+ and Fe3+. In a 2-h incubation with laminaridextrins (seven to nine glucose units) or curdlan and excess enzyme, the major products were glucose (30 to 37%), laminaribiose (17 to 23%), laminaritriose (18 to 28%), laminaritetraose (13 to 21%), and small amounts of large laminarioligosaccharides. With laminarihexaose and laminaripentaose, the products were equal quantities of laminaribiose and glucose (30%) and laminaritetraose and laminaritriose (18 to 21%). Laminaribiose or laminaritriose were not hydrolyzed, indicating a requirement for at least four contiguous 1,3-beta-linked glucose units for enzyme activity. The enzyme appeared to have the properties of both an exo- and an endoglucanase.(ABSTRACT TRUNCATED AT 250 WORDS)     

丽斗蟋翅二型雄虫食物消化能力及消化酶活性比较

【目的】丽斗蟋Velarifictorus ornatus具明显的翅二型现象,为探讨翅型分化对丽斗蟋翅二型雄虫消化能力及中肠内消化酶活性产生的影响,对长翅型与短翅型雄虫食物消化能力及中肠内消化酶活性进行了检测比较。【方法】我们采取重量营养指数测定了羽化后12 d内丽斗蟋两型雄成虫增长量、相对增长率、取食量、食物利用率、近似消化率和食物转化率。为进一步明确丽斗蟋翅二型成虫食物消化能力与中肠内消化酶活性的关系,我们采用4种专用底物测定了中肠内用于分解蛋白质、脂肪和碳水化合物的总蛋白酶、胰蛋白酶、脂肪酶和淀粉酶的活性。【结果】结果表明,丽斗蟋两型雄虫取食量、食物转化率、食物利用率与增长量均无统计差异,但中肠内消化酶活性变化规律不同。成虫羽化后4 d时,长翅型雄虫中肠内总蛋白酶与胰蛋白酶活性显著高于短翅型雄虫,相反,羽化后0 d时,短翅型雄虫中肠内总蛋白酶与胰蛋白酶活性则显著高于长翅型雄虫,而羽化后12 d时,虽然短翅型雄虫总蛋白酶活性高于长翅型雄虫,但胰蛋白酶活性在两型雄虫间并无差异。成虫羽化后0 d时,两型雄虫脂肪酶活性无差异,但无论是羽化后4 或 12 d,长翅型雄虫中肠内脂肪酶活性皆显著大于短翅型雄虫。成虫羽化后4 d时,短翅型雄虫中肠内淀粉酶活性显著高于长翅型雄虫,而羽化后0与12 d时,两型雄虫间无显著差异。【结论】丽斗蟋翅二型雄虫食物消化能力无显著差异,但羽化后不同时间,中肠内消化酶活性存在差异,该差异可能与成虫羽化后不同时期,翅二型雄虫在飞行与繁殖投资中对不同能源物质的需求有关。     

The effect of acquired microbial enzymes on assimilation efficiency in the common woodlouse,Tracheoniscus rathkei

Summary The digestive tract of the common woodlouse, Tracheoniscus rathkei Brandt (Isopoda: Oniscoidea), contains digestive enzymes active against -1,4-glucans, which are the chief storage polysaccharides of vascular plants, algae, fungi, and animals, and -1,3-glucans, which are present in algae and fungi. Digestive tract extracts also exhibit significant activity toward xylan and carboxymethyl-cellulose but negligible activity toward microcrystalline cellulose, substrates representative of the major structural polysaccharides of vascular plants. Low activity was detected toward pectin, and no activity was detected toward chitin. Activity toward xylan is due in part to microbial enzymes acquired from the leaf litter which was the isopod's normal food. Although ingested microbial xylanases are stable and active in the gut fluid, they do not make a quantitatively significant contribution to the isopod's ability to assimilate the hemicellulosic component of its diet. However, the assimilation of carbon from labeled plant fiber is enhanced in isopods which have acquired a cellulase by ingestion of leaf litter amended with a commercial preparation of the cellulase complex from the fungus, Penicillium funiculosum. This result demonstrates the potential contribution of acquired enzymes to the digestion of plant fiber in terrestrial detritivores. We urge caution, however, in assigning an important digestive function to ingested enzymes on the basis of evidence that only indicates that such enzymes are present in the gut fluid without additional evidence that their presence results in an enhancement of digestive efficiency.     

The digestive gland of the Southern Dumpling Squid (Euprymna tasmanica): structure and function

The cephalopod digestive gland plays an important role in the efficient assimilation of nutrients and therefore the fast growth of the animal. The histological and enzymatic structure of Euprymna tasmanica was studied and used in this experiment to determine the dynamics of the gland in response to feeding. The major roles of the digestive gland were secretion of digestive enzymes in spherical inclusions (boules) and excretion of metabolic wastes in brown body vacuoles. High levels of trypsin, chymotrypsin and α-amylase, low levels of α-glucosidase and negligible carboxypeptidase activity were produced by the gland. There was no evidence of secretion of digestive enzymes in other organs of the digestive tract. Within 60 min of a feeding event, the gland produced increasing numbers of boules to replace those lost from the stomach during the feeding event. Initially, small boules were seen in the digestive cells, they increased in size until they are released into the lumen of the gland where they are transported to the stomach. There was no evidence of an increase in activity of digestive enzymes following a feeding event, despite structural changes in the gland. However, there was large variation among individuals in the level of digestive enzyme activity. A negative correlation between boule and brown body vacuole density suggested that the large variation in enzyme activity may be due to the digestive gland alternating between enzyme production and excretion.     

Isolation and properties of a (1,3)-beta-D-glucanase from Ruminococcus flavefaciens.

A (1,3)-beta-D-glucanase [(1,3)-beta-D-glucan-3-glucanohydrolase] from Ruminococcus flavefaciens grown on milled filter paper was purified 3,700-fold (19% yield) and appeared as a single major protein and activity band upon polyacrylamide gel electrophoresis. The enzyme did not hydrolyze 1,6-beta linkages (pustulan) or 1,3-beta linkages in glucans with frequent 1,6-beta-linkage branch points (scleroglucan). Curdlan and carboxymethylpachyman were hydrolyzed at 50% the rate of laminarin. The enzyme had a Km of 0.37 mg of laminarin per ml, a pH optimum of 6.8, and a temperature optimum of 55 degrees C and was stable to heating at 40 degrees C for 60 min. The molecular mass of the enzyme was estimated to be 26 kDa by gel filtration and 25 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme was completely inhibited by 1 mM Hg2+, Cu2+, and KMnO4, 75% by 1 mM Ag2+, and Ni2+, and 50% by 1 mM Mn2+ and Fe3+. In a 2-h incubation with laminaridextrins (seven to nine glucose units) or curdlan and excess enzyme, the major products were glucose (30 to 37%), laminaribiose (17 to 23%), laminaritriose (18 to 28%), laminaritetraose (13 to 21%), and small amounts of large laminarioligosaccharides. With laminarihexaose and laminaripentaose, the products were equal quantities of laminaribiose and glucose (30%) and laminaritetraose and laminaritriose (18 to 21%). Laminaribiose or laminaritriose were not hydrolyzed, indicating a requirement for at least four contiguous 1,3-beta-linked glucose units for enzyme activity. The enzyme appeared to have the properties of both an exo- and an endoglucanase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation and characterization of two types of β-1,3-glucanases from the common sea hare Aplysia kurodai

Two types of β-1,3-glucanases, AkLam36 and AkLam33 with the molecular masses of 36 kDa and 33 kDa, respectively, were isolated from the digestive fluid of the common sea hare Aplysia kurodai. AkLam36 was regarded as an endolytic enzyme (EC 3.2.1.6) degrading laminarin and laminarioligosaccharides to laminaritriose, laminaribiose, and glucose, while AkLam33 was regarded as an exolytic enzyme (EC 3.2.1.58) directly producing glucose from polymer laminarin. AkLam36 showed higher activity toward β-1,3-glucans with a few β-1,6-linked glucose branches such as Laminaria digitata laminarin (LLam) than highly branched β-1,3-glucans such as Eisenia bicyclis laminarin (ELam). AkLam33 showed moderate activity toward both ELam and LLam and high activity toward smaller substrates such as laminaritetraose and laminaritriose. Although both enzymes did not degrade laminaribiose as a sole substrate, they were capable of degrading it via transglycosylation reaction with laminaritriose. The N-terminal amino-acid sequences of AkLam36 and AkLam33 indicated that both enzymes belong to the glycosyl hydrolase family 16 like other molluscan β-1,3-glucanases.     

Digestive enzymes of two brachyuran and two anomuran land crabs from Christmas Island,Indian Ocean

The digestive ability of four sympatric land crabs species (the gecarcinids, Gecarcoidea natalis and Discoplax celeste and the anomurans, Birgus latro and Coenobita perlatus) was examined by determining the activity of their digestive enzymes. The gecarcinids are detritivores that consume mainly leaf litter; the robber crab, B. latro, is an omnivore that preferentially consumes items high in lipid, carbohydrate and/or protein; C. perlatus is also an omnivore/detritivore. All species possess protease, lipase and amylase activity for hydrolysing ubiquitous protein, lipid and storage polysaccharides (glycogen and starch). Similarly all species possess enzymes such as N-acetyl-β-d-glucosaminidase, the cellulases, endo-β-1,4-glucanase and β-glucohydrolase and hemicellulases, lichenase and laminarinase for the respective hydrolysis of structural substrates chitin, cellulose and hemicelluloses, lichenan and laminarin. Except for the enzyme activities of C. perlatus, enzyme activity could not be correlated to dietary preference. Perhaps others factors such as olfactory and locomotor ability and metabolic status may determine the observed dietary preferences. The digestive fluid of C. perlatus possessed higher endo-β-1,4-glucanase, lichenase and laminarinase activities compared to that of the other species. Thus, C. perlatus may be efficient at digestion of cellulose and hemicellulose within plant material. Zymography indicated that the majority of protease, lipase, phosphatase, amylase, endo-β-1,4-glucanase, β-glucohydrolase and N-acetyl-β-d-glucosaminidase isozymes were common to all species, and hence were inherited from a common aquatic ancestor. Differences were observed for the phosphatase, lipase and endo-β-1,4-glucanase isozymes. These differences are discussed in relation to phylogeny and possible evolution to cope with the adoption of a terrestrial diet.     

Cellulase Complex of the Fungus Chrysosporium lucknowense: Isolation and Characterization of Endoglucanases and Cellobiohydrolases

Using different chromatographic techniques, eight cellulolytic enzymes were isolated from the culture broth of a mutant strain of Chrysosporium lucknowense: six endoglucanases (EG: 25 kD, pI 4.0; 28 kD, pI 5.7; 44 kD, pI 6.0; 47 kD, pI 5.7; 51 kD, pI 4.8; 60 kD, pI 3.7) and two cellobiohydrolases (CBH I, 65 kD, pI 4.5; CBH II, 42 kD, pI 4.2). Some of the isolated cellulases were classified into known families of glycoside hydrolases: Cel6A (CBH II), Cel7A (CBH I), Cel12A (EG28), Cel45A (EG25). It was shown that EG44 and EG51 are two different forms of one enzyme. EG44 seems to be a catalytic module of an intact EG51 without a cellulose-binding module. All the enzymes had pH optimum of activity in the acidic range (at pH 4.5-6.0), whereas EG25 and EG47 retained 55-60% of the maximum activity at pH 8.5. Substrate specificity of the purified cellulases against carboxymethylcellulose (CMC), beta-glucan, Avicel, xylan, xyloglucan, laminarin, and p-nitrophenyl-beta-D-cellobioside was studied. EG44 and EG51 were characterized by the highest CMCase activity (59 and 52 U/mg protein). EG28 had the lowest CMCase activity (11 U/mg) amongst the endoglucanases; however, this enzyme displayed the highest activity against beta-glucan (125 U/mg). Only EG51 and CBH I were characterized by high adsorption ability on Avicel cellulose (98-99%). Kinetics of Avicel hydrolysis by the isolated cellulases in the presence of purified beta-glucosidase from Aspergillus japonicus was studied. The hydrolytic efficiency of cellulases (estimated as glucose yield after a 7-day reaction) decreased in the following order: CBH I, EG60, CBH II, EG51, EG47, EG25, EG28, EG44.     

Lysis of yeast cell walls. Lytic beta-(1 leads to 3)-glucanases from Bacillus circulans WL-12.

Bacillus circulans WL-12 when grown in a mineral medium with yeast cell walls or yeast glucan as the soli carbon source, produced five beta-glucanases. Two beta-(1 leads to 3)-glucanases (I and II), which are lytic to yeast cell walls, were isolated from the culture liquid by batch adsorption on yeast glucan, and separated by chromatography on hydroxylapatite. Lytic beta-(1 leads to 3)-glucanase I was further purified by carboxymethylcellulose chromatography. The specific activity of lytic beta-(1 leads to 3)-glucanase I on laminarin was 4.1 U per mg of protein. The enzyme moved as a single protein with a molecular weight of 40000 during sodium dodecylsulfate electrophoresis in slab gels. It was specific for the beta-(1 leads to 3)-glucosidic bond but the enzyme did not hydrolyze laminaribiose. Hydrolysis of laminarin went through a series of oligosaccharides, and laminaribiose and glucose accumulated till the end of the reaction. A small amount of gentibiose was also produced from laminarin. Products from yeast cell walls and yeast glucan included laminaripentaose, laminaritriose, laminaribiose, glucose and gentiobiose, but no laminaritetraose was detected. This glucanase has an optimum pH of 5.5.     

Purification and Characterization of an Endo-(1,3)-beta-d-Glucanase from Trichoderma longibrachiatum

A laminarinase [endo-(1,3)-beta-d-glucanase] has been purified from Trichoderma longibrachiatum cultivated with d-glucose as the growth substrate. The enzyme was found to hydrolyze laminarin to oligosaccharides varying in size from glucose to pentaose and to lesser amounts of larger oligosaccharides. The enzyme was unable to cleave laminaribiose but hydrolyzed triose to laminaribiose and glucose. The enzyme cleaved laminaritetraose, yielding laminaritriose, laminaribiose, and glucose, and similarly cleaved laminaripentaose, yielding laminaritetraose, laminaritriose, laminaribiose, and glucose. The enzyme cleaved only glucans containing beta-1,3 linkages. The pH and temperature optima were 4.8 and 55 degrees C, respectively. Stability in the absence of a substrate was observed at temperatures up to 50 degrees C and at pH values between 4.9 and 9.3. The molecular mass was determined to be 70 kilodaltons by sodium dodecyl sulfate-12.5% polyacrylamide gel electrophoresis, and the pI was 7.2. Enzyme activity was significantly inhibited in the presence of HgCl(2), MnCl(2), KMnO(4), and N-bromosuccinimide. The K(m) of the enzyme on laminarin was 0.0016%, and the V(max) on laminarin was 3,170 mumol of glucose equivalents per mg of the pure enzyme per min.     

Comparative studies on beta-glucan hydrolases. Isolation and characterization of an exo(1 yields 3)-beta-glucanase from the snail, Helix pomatia.

An exo-β-glucan hydrolase, present in the digestive juice of the snail, Helix pomatia, has been purified to homogeneity by chromatography on Bio-Gel P-60, Sephadex G-200, DEAE-cellulose, and DEAE-Sephadex. The enzyme degrades β-(1 → 3)-linked oligosaccharides and polysaccharides, rapidly and to completion, or near completion, yielding glucose as the major product of enzyme action. Mixed linkage (1→3; 1→4)-β-glucans are also extensively degraded and β-(1→6)- and β-(1→4)-linked glucose polymers are slowly degraded by the enzyme. This enzyme differs from other exo-β-glucanases, reported previously, in the broadness of its substrate specificity. The K_m values for action on laminarin and lichenin are respectively 1.22 and 2.22 mg/ml; the maximum velocity of action on laminarin is approximately twice that on lichenin. The enzyme has a molecular weight of 82,000 as determined by polyacrylamide gel electrophoresis. Maximum activity is exhibited at pH 4.3 and at temperatures of 50–55 °C.     

Digestive capabilities reflect the major food sources in three species of talitrid amphipods

Digestive enzyme activities of three talitrid amphipods were examined to investigate the relationship between their digestive capabilities and diet. Laminarinase, cellobiase, carboxymethyl-cellulase, xylanase, alpha- and beta-glucosidase and lipase were detected in all three species suggesting talitrid amphipods can readily digest dietary carbohydrate and lipid, including complex polysaccharides. Relatively high specific enzyme activity (Units (mg(-1) digestive tract protein)(-1)) of laminarinase and lipase was detected in Talorchestia marmorata, a supralittoral kelp feeder which is coherent with the digestion of lipid-esters and beta-glucans (laminarin) which are the main lipid and storage polysaccharides of brown seaweeds. Talorchestia sp., a low shore intertidal feeder, had high enzymatic activity of alpha- and beta-glucosidase, cellobiase and xylanase, which is consistent with the digestion of diatoms. Keratroides vulgaris, a forest litter feeder had a relatively low specific activity of all enzymes. It is possible that leaf litter is partially digested prior to ingestion by bacteria and fungi present in the rotting vegetation, with bacterial and fungal enzymes contributing to this species' ability to hydrolyse its diet. This study provides the first quantitative data on digestive capacity in these three talitrid amphipods and confirms the relationship between dietary preference and digestive enzyme complement.     

The extracellular enzymes of Coniophora cerebella

1. The extracellular enzymes present in the culture filtrates of Coniophora cerebella grown on various carbohydrate carbon sources were investigated. Enzymes that degraded cellulose derivatives, hemicellulose fractions, starch, laminarin, pectin and several oligosaccharides were detected. 2. All the polysaccharide-degrading activities were adaptive except for that acting on laminarin. 3. The culture filtrates degraded native cellulose to only a very limited extent. 4. The hemicellulase activity included enzymes acting on all the major components of wood hemicellulose. 5. The main starch-degrading enzyme was a glucoamylase. 6. Laminarin-degrading activity was produced when cellulose, hemicellulose or starch was used as carbon source for the fungus and it may be involved in the re-utilization of hyphal carbon or of a reserve polysaccharide synthesized during active growth of the organism.