Mol Hexose Research Articles

Characterization of the cellulolytic and hydrogen-producing activities of six mesophilic Clostridium species

To characterize cellulolytic, hydrogen-producing clostridia on a comparable basis. H(2) production from cellulose by six mesophilic clostridia was characterized in standardized batch experiments using MN301 cellulose, Avicel and cellobiose. Daily H(2) production, substrate degradation, biomass production and the end-point distribution of soluble fermentation products varied with species and substrates. All species produced a significant amount of H(2) from cellobiose, with Clostridium acetobutylicum achieving the highest H(2) yield of 2.3 mol H(2) mol(-1) hexose, but it did not degrade cellulose. Clostridium cellulolyticum and Clostridium populeti catalysed the highest H(2) production from cellulose, with yields of 1.7 and 1.6 mol H(2 )mol(-1) hexose from MN301 and 1.6 and 1.4 mol H(2) mol(-1) hexose from Avicel, respectively. These species also achieved 25-100% higher H(2) production rates from cellulose than the other species. These cellulolytic, hydrogen-producing clostridia varied in H(2) production, with Cl. cellulolyticum and Cl. populeti achieving the highest H(2) yields and cellulose degradation. The fermentation of cellulosic materials presents a means of H(2) production from renewable resources. This standardized comparison provides a quantitative baseline for improving H(2) production from cellulose through medium and process optimization and metabolic engineering.

Simultaneous production of biohydrogen and bioethanol with fluidized-bed and packed-bed bioreactors containing immobilized anaerobic sludge

Hydrogen and ethanol are promising biofuels and of great potential to become alternatives to fossil fuels. In this work, two bioreactor systems, namely fluidized-bed (FBR) and packed-bed (PBR), were developed to produce H 2 and ethanol simultaneously from dark fermentation of carbohydrate substrates using polyethylene–octane elastomer immobilized anaerobic sludge as the biocatalyst. The H 2 and ethanol production in FBR essentially increased with increasing upflow velocity ( v up ), as sucrose and fructose was better substrate for the yield of H 2 and ethanol, respectively. With FBR operated at v up = 0.91 cm s − 1 , sucrose gave the highest H 2 production rate (59 mmol h −1 l −1) among the three sugar substrates (sucrose, glucose, and fructose) tested, but the best H 2 yield (1.04 mol mol hexose −1) was obtained with glucose at v up = 0.55 cm s − 1 . For ethanol production in FBR, fructose was the favorable substrate, resulting in maximum ethanol production rate and yield of 378 mmol h −1 l −1 and 0.65 mol mol hexose −1, respectively, when operating at v up = 0.91 cm s − 1 . At a hydraulic retention time of 4 h, the PBR system produced H 2 and ethanol at a slower rate of 16 and 6 mmol h −1 l −1, respectively, by using glucose. However, the yields of H 2 and ethanol were comparable to those for FBR. The soluble metabolites were dominated by ethanol, accounting for 43–65% of total soluble microbial products. The production of acetate and butyrate was less significant when compared to cultures optimized for H 2 production. Comparison of the yield of H 2 and ethanol shows that production of H 2 and ethanol was reversely correlated. The total energy generation based on the heat values of H 2 and ethanol was calculated to assess the overall efficiency of energy production. In FBR, the energy generation rate was higher when a faster upflow velocity was used. The best energy generation rate and yield was 526 kJ h −1 l −1 and 1048 kJ mol hexose −1, respectively, both occurred with fructose-feeding FBR operated at v up = 0.91 cm s − 1 . The PBR system displayed a lower energy generation rate, whereas the energy yield was comparable or even higher than those for FBR.

Performance characteristics of a two‐stage dark fermentative system producing hydrogen and methane continuously

The performance of a mesophilic two-stage system generating hydrogen and methane continuously from sucrose (10-30 g/L) was investigated. A hydrogen-generating CSTR followed by an upflow anaerobic filter were both inoculated with anaerobically digested sewage sludge, and ORP, pH, gas output, %H(2), %CH(4) and %CO(2) monitored. pH was controlled with NaOH, KOH or Ca(OH)(2). Using NaOH as alkali with 10 g/L sucrose, yields of 1.62 +/- 0.2 mol H(2)/mol hexose added and 323 mL CH(4)/gCOD added to the hydrogen and methane reactors respectively were achieved. The overall chemical oxygen demand (COD) reduction was 92.6% with 0.90 +/- 0.1 g/L sodium and 316 +/- 40 mg/L residual acetate in the methane reactor. Operation at 20 g/L sucrose and NaOH as alkali led to impaired volatile fatty acid (VFA) degradation in the methane reactor with 2.23 +/- 0.2 g/L sodium, 1,885 mg/L residual acetate, a hydrogen yield of 1.47 +/- 0.1 mol/mol hexose added, a methane yield of 294 mL/gCOD added and an overall COD reduction of 83%. Using Ca(OH)(2) as alkali with 20 g/L sucrose gave a hydrogen yield of 1.29 +/- 0.3 mol/mol hexose added, a methane yield of 337 mL/gCOD added and improved the overall COD reduction to 91% with residual acetate concentrations of 522 +/- 87 mg/L. Operation at 30 g/L sucrose with Ca(OH)(2) gave poorer overall COD reduction (68%), a hydrogen yield of 1.47 +/- 0.2 mol/mol hexose added, a methane yield of 138 mL/gCOD added and residual acetate 7,343 +/- 715 mg/L. It was shown that sodium toxicity and overloading are important issues for successful anaerobic digestion of effluent from biohydrogen reactors in high rate systems.

Continuous hydrogen production by anaerobic mixed microflora using a hollow-fiber microfiltration membrane bioreactor

A membrane bioreactor (MBR) fabricated by connecting a hollow-fiber microfiltration membrane module with a continuous flow stirred tank reactor (CSTR) was used to enhance H 2 production through high-dilution rate operations. Three different carbon substrates (glucose, sucrose, and fructose) were examined for their effectiveness in H 2 production with a mixed microflora. The results show that in CSTR operation, cell washout occurred at a hydraulic retention time (HRT) of 2–4 h. Using MBR could avoid cell washout, leading to a substantial increase in both H 2 production rate (HPR) and biomass concentration. The MBR system was very effective in retaining biomass within the reactor as the system can be stably operated at an extremely low HRT of 1 h with an optimal steady-state HPR of 1.48, 2.07, and 2.75 l/h/l, respectively, for using glucose, sucrose, and fructose as the sole carbon source. Meanwhile, despite operation at a high dilution rate (i.e., HRT = 1 h ), the H 2 yield (HY) could be maintained at a high level of 1.27, 1.39, and 1.36 mol H 2 / mol hexose, for glucose, sucrose, and fructose, respectively. Irrespective of the bioreactor mode (CSTR or MBR), the HPR tended to decrease in the order of fructose > sucrose > glucose . Thus, fructose seems to be the most efficient carbon substrate for H 2 production with the H 2 -producing mixed culture used in this work. Butyrate (HBu) and acetate (HAc) (especially, HBu) were the major soluble metabolites in all cases, contributing to 70–85% of total soluble microbial product (SMP). The HPR and HY could be estimated based on stoichiometric correlation between formation of soluble metabolites (i.e., HBu, HAc, and propionate) and H 2 production. The estimated values are in good agreement with the experimental results.

Effect of gas sparging on continuous fermentative hydrogen production

The effect of gas sparging on continuous fermentative H 2 production was investigated in completely stirred-tank reactors (CSTR) using internal biogas, N 2 and CO 2 with various flow rates (100, 200, 300 and 400 ml/min). The sparging with external gases of N 2 and CO 2 showed higher H 2 yield than the control of no sparging and internal biogas sparging. It indicated that the decrease of H 2 partial pressure by external gas sparging had a beneficial effect on H 2 fermentation. Especially, CO 2 sparging was more effective in the reactor performance than N 2 sparging, accompanied by higher production of H 2 and butyrate. The best performance was obtained by CO 2 sparging at 300 ml/min, resulting in the highest H 2 yield of 1.68 mol H 2 / mol hexose consumed and the maximum specific H 2 production rate of 6.89 L H 2 / g VSS/day. Compared to N 2 sparging, there might be another positive effect in CO 2 sparging apart from lowering H 2 partial pressure. High CO 2 partial pressure had little effect on H 2 -producing bacteria but inhibitory effect on other microorganisms such as acetogens and lactic acid bacteria which were competitive with H 2 -producing bacteria. Only H 2 -producng bacteria, such as Clostridium tyrobutyricum, C. proteolyticum and C. acidisoli were isolated under CO 2 sparging conditions based on 16S rDNA analysis by PCR-DGGE.

Continuous bio-hydrogen production from citric acid wastewater via facultative anaerobic bacteria

In this study, continuous biological hydrogen production using wastewater from citric acid factory as raw materials was investigated. And enrichment cultures from facultative hydrogen-producing anaerobe together with citric acid wastewater were utilized. For finding out the industrialized feasibility of continuous H 2 bio-production, the ability of H 2 -production via facultative anaerobe, optimum hydraulic retention time (HRT) and optimum volume loading rate (VLR) were also studied. The experimental results showed that facultative anaerobic bacteria could yield hydrogen as high as 0.84 mol H 2 / mol hexose consumed at the VLR of 38.4 kg COD / m 3 d . Butyric acid was the dominant product among volatile fatty acids (VFAs). An improved upward-flow anaerobic sludge blanket (UASB) reactor with a working volume 50 m 3 was used. Continuous hydrogen yield was 0.72 m 3 H 2 / m 3 reactor d under the following conditions: temperature at 35– 38 ∘ C , initial pH at 6.8–7.2, HRT = 12 h . The study also indicates that the reactor used has a better operational stability and the facultative anaerobe has an excellent adaptive capacity for organic loading rate. Furthermore, the chemical oxygen demand (COD) removal efficiency exceeded 60% and the total sugar degradation efficiency (TSDE) was close to or over 90%, even up to 96.6% at HRT of 12 h during the experiment.

Continuous fermentative hydrogen production from sucrose and sugarbeet

To produce hydrogen by fermentation of biomass, a continuous process using a non-sterile substrate with a readily available mixed microflora is desirable. This work investigates a simple batch start-up procedure at pH 5.2 and 32°C, using anaerobically digested sewage sludge, and continuous hydrogen production from refined sucrose, pulped sugarbeet and a water extract of sugarbeet. Without heat treating the sludge, and with initial nitrogen sparging, a hydrogen producing culture was established within 5 days and remained stable during two experiments of 45 and 32 days duration. At 14– 15 h retention time ( 16 kg total sugar m −3 d −1 organic loading rate) hydrogen yields for refined sucrose and pulped sugarbeet were, respectively, 1.0±0.1 and 0.9±0.2 mol/ mol hexose converted. With nitrogen sparging hydrogen yields were 1.7±0.2–1.9±0.2 and 1.7±0.2 mol /mol hexose converted for refined sucrose and water extract of sugarbeet, respectively. Increasing ethanol concentration during operation on sugarbeet, and in some cases a higher redox potential (>−150 mV) , correlated with lower hydrogen yield.

Characterisation of a thermostable pepstatin-insensitive acid proteinase from a Bacillus sp.

An acid proteinase, Wai 21a, produced by a thermophilic Bacillus species (strain Wai 21a) has been purified to homogeneity by cation-exchange chromatography, phenyl-Sepharose chromatography and anion-exchange chromatography. A pI of 3.8 was determined by isoelectric focussing. The protein contained some associated carbohydrate (20 mol hexose equiv/mol proteinase). Optimal proteolytic activity was observed at pH 3.0 (at 60°C). The Leu 15-Tyr 16 bond was the major site of hydrolysis for the oxidized B chain of insulin. Enzyme activity was not affected by inhibitors of the cysteine, metallo or serine class of proteinases. The aspartate proteinase inhibitor, pepstatin, did not inhibit enzyme activity. Inhibition of enzyme activity by 1,2-epoxy-3-( p-nitrophenoxy)-propane indicated the presence of at least one carboxyl group essential to the catalytic mechanism of the enzyme. Proteinase activity was inhibited by diazoacetyl- dl-norleucine methyl ester in a slow and non-specific manner atypical of pepstatin-sensitive aspartate proteinases. Wai 21a proteinase may be classified as member of the pepstatin-insensitive group of aspartate proteinases. The thermal stability at pH 3.0 and 60°C increased 2.1-fold ( t 1 2 , 4.5–9.7 hr) in the presence of 5 mM Ca ++. An increase in both pH (3.0–4.5) and Ca ++ concentration (0–30 mM) resulted in a 15-fold increase ( t 1 2 , 15–230 min) in thermal stability at 75°C. The amino acid composition of Wai 21a proteinase was found to be similar to other pepstatin-insensitive proteinases from bacterial sources and in particular similar to the thermostable enzyme, kumamolysin.

Identification of a Disaccharide (Xyl-Glc) and a Trisaccharide (Xyl2-Glc) O-Glycosidically Linked to a Serine Residue in the First Epidermal Growth Factor-like Domain of Human Factors VII and IX and Protein Z and Bovine Protein Z

We have recently described a unique trisaccharide linked to a serine residue in the first epidermal growth factor-like domains of bovine blood coagulation factors VII (Ser-52) and IX (Ser-53) (Hase, S., Kawabata, S., Nishimura, H., Takeya, H., Sueyoshi, T., Miyata, T., Iwanaga, S., Takao, T., Shimonishi, Y., and Ikenaka, T. (1988) J. Biochem. (Tokyo) 104, 867-868). The sugar chain identified in these clotting factors consists of 1 mol of hexose (glucose (Glc] and 2 mol of pentose (xylose (Xyl]. We report here that human factors VII and IX and protein Z and bovine protein Z also contain such carbohydrate moieties linked to a serine residue at the same position found in bovine factors VII and IX. A glycopeptide derived from each of these proteins was subjected to amino acid sequence and component sugar analyses and fast atom bombardment mass spectrometric analysis. The results indicate that the glycopeptide derived from human factor IX contains 1 mol each of Glc and Xyl. The reducing end of this disaccharide was identified as Glc by analyzing the disaccharide generated by hydrazinolysis. In contrast, human factor VII and protein Z yielded two different glycopeptides which contained Glc and Xyl at molar ratios of 1:1 and 1:2, respectively, suggesting microheterogeneity of these O-linked sugar chains. Bovine protein Z glycopeptide contained 1 mol of Glc and 2 mol of Xyl. These sugar compositions were confirmed by analyses of the intact proteins. In relation to the trisaccharide sugar chain previously discovered in bovine factors VII and IX, these findings indicate the existence of a Xyl2-Glc-Ser and a Xyl-Glc-Ser structure in the first epidermal growth factor-like domains of human factors VII and IX and protein Z in addition to that of bovine protein Z. Whether these carbohydrate moieties contribute to the biological activities of these proteins is unknown.

Isolation and characterization of soluble β-galactoside-binding lectins from mammalian liver

Soluble β-galactoside-binding lectins were isolated by chromatography on asialofetuin-Sepharose-4B column in 10 mM Tris-HCl buffer (pH 7.5) containing 150 mM NaCl, 5 mM CaCl 2 and 1 mM 2-mercaptoethanol. The three lectins moved essentially as single polypeptide bands, of 18, 22 and 24 kDa, respectively, for sheep, goat and buffalo hepatic lectins. Sheep and goat lectins each contained 4 mol of hexose, whereas the hexose content of the buffalo lectin was 7 mol. The number of sulfhydryl groups in sheep, goat and buffalo lectins were determined to be 3.2, 4.3 and 4.8, respectively. The optical properties of the three lectins were similar to those of tryptophan-containing proteins. Lectin mediated hemagglutination of trypsinized rabbit erythrocytes was most effectively inhibited by lactose, followed by o-nitrophenyl β-galactopyranoside and galactose, but remained unaffected by glucose, mannose, fucose and fructose. Calcium ions substantially enhanced their hemagglutinating activity. Goat and buffalo lectins, but not sheep lectin, were also stimulated by Mg 2+, Mn 2+, Sr 2+ and Ni 2+ ions. The lectins lost activity after treatment with para-hydroxy-mercuribenzoate and N-ethylmaleimide. However, iodoacetamide treatment had no effect on the activity. The results show that the three lectins are different from the soluble β-galactoside-binding lectins studied thus far.

Isolation and characterization of cancer procoagulant: a cysteine proteinase from malignant tissue.

Cancer procoagulant, a proteolytic procoagulant enzyme, has been purified from rabbit V2 carcinoma extracts by two procedures. In the first, the protein was purified by benzamidine--Sepharose affinity chromatography, gel filtration chromatography, and phenyl-Sepharose hydrophobic chromatography. Antiserum was raised against the purified protein and was used to prepare an immunoadsorbent column. In the second, tumor extracts were purified by immunoaffinity chromatography followed by p-(chloromercuri)benzoate affinity chromatography. The second procedure was substantially quicker and easier. The final product of both procedures was homogeneous on the basis of analytical sodium dodecyl sulfate--polyacrylamide gel electrophoresis and isoelectric focusing. The molecular weight was 68 000 and the isoelectric point 4.8. The proteinase activity of cancer procoagulant directly activated factor X, in the absence of factor VII, and was inhibited by 1 mM iodoacetamide and 0.1 mM mercury which are classic cysteine proteinase inhibitors. A carbohydrate analysis showed less than 1 mol of hexose or sialic acid/mol of protein. The amino acid analysis showed that serine (19.1%), glycine (18.77%), and glutamic acid (12.5%) were the prevalent amino acids. The amino acid composition of cancer procoagulant was substantially different than other known factor X activating proteinases or other cysteine proteinases including cathepsin B.

Acetomicrobium flavidum, gen. nov., sp. nov., a thermophilic, anaerobic bacterium from sewage sludge, forming acetate, CO 2 and H 2 from Glucose

Summary Acetomicrobium flavidum gen. nov., sp. nov. was isolated from a thermophilic, anaerobic sewage sludge digester operated at 60 °c. The Gram-negative, highly motile, non-sporeforming rods form 2 mol acetic acid, 2 mol CO 2 and 4 mol H 2 from 1 mol hexose. They represent at least 25% of the total population (microscopic count) of the fermenting sludge.

Fermentation of Cellulose to Methane and Carbon Dioxide by a Rumen Anaerobic Fungus in a Triculture with Methanobrevibacter sp. Strain RA1 and Methanosarcina barkeri

The fermentation of cellulose by a rumen anaerobic fungus in the presence of Methanobrevibacter sp. strain RA1 and Methanosarcina barkeri strain 227 resulted in the formation of 2 mol each of methane and carbon dioxide per mol of hexose fermented. Coculture of the fungus with either Methanobrevibacter sp. or M. barkeri produced 0.6 and 1.3 mol of methane per mol of hexose, respectively. Acetate, formate, ethanol, hydrogen, and lactate, which are major end products of cellulose fermentation by the fungus alone, were either absent or present in very low quantities at the end of the triculture fermentation (</=0.08 mol per mol of hexose fermented). During the time course of cellulose fermentation by the triculture, hydrogen was not detected (<1 x 10 atm; <0.001 kPa) and only acetate exhibited transitory accumulation; the maximum was equivalent to 1.4 mol per mol of hexose at 6 days which was higher than the total acetate yield of 0.73 in the fungus monoculture. The effect of methanogens is interpreted as a shift in the flow of electrons away from the formation of electron sink products lactate and ethanol to methane via hydrogen, favoring an increase in acetate, which is in turn converted to methane and carbon dioxide by M. barkeri. The maximum rate of cellulose degradation in the triculture (3 mg/ml per day) was faster than previously reported for bacterial cocultures and within 16 days degradation was complete. The triculture was used successfully also in the production of methane from cellulose in the plant fibrous materials, sisal (fiber from leaves of Agave sisalona L.) and barley straw leaf.

Secretion of endogenous lectin by chicken intestinal goblet cells.

The two lactose-binding lectins found in adult chicken intestine, chicken-lactose-lectin-1 (CLL-1) and chicken-lactose-lectin-11 (CLL-11), were localized within the vesicles of the mucin-secreting goblet cells by indirect immunofluorescence and immunoperoxidase staining methods. Attention was concentrated on CLL-11 which is 200 time more abundant than CLL-1 in adult intestine. The localization of CLL-11 in secretory vesicles, combined with its demonstration on the intestinal epithelial surface by immune staining methods and by specific elution with lactose, suggested that at least a portion of the CLL-11 in the vesicles was secreted by the goblet cells and then became associated with the mucosal surface. In support of this, treatment of isolated intestinal strips with a cholinergic agent, bethanechol (10(-7 M) produced a small but significant increase in the amount of CLL-11 that could be eluted from their surface with lactose. Secretion of lectin may occur in conjunction with mucin because both are localized in the secretory vesicles and CLL-1 and CLL-11 apparently bind to purified chicken intestinal mucin, which is a potent inhibitor of their hemagglutination activities. The mucin is six orders of magnitude more potent than lactose as a hemagglutination inhibitor of CLL-1 or CLL-11 on a molar basis, and three orders of magnitude more potent when expressed per mole of hexose. These results suggest that CLL-11, and perhaps CLL-1, are secreted from the goblet cells along with mucin. They may function in the organization of mucin for secretion and/or in its association with the intestinal mucosal surface.

Characterization of the hemoglobin of Biomphalaria glabrata as a glycoprotein

1. 1. The hemoglobin of Biomphalaria glabrata is a glycoprotein which contains 3% sugars. 2. 2. The sugar moiety is composed of 2 mol of hexose and one mol of hexosamine per mol of monomer. 3. 3. The hexosamine in the molecule is glucosamine. 4. 4. Mannose, galactose and fucose are present in the molecule in a ratio of 2:1:1. 5. 5. No sialic and uronic acids were detected in this hemoglobin.

Protective role of increased myocardial glycogen stores in cardiac anoxia in the rat.

To determine whether increased glycogen stores might protect the heart against anoxia, experiments were performed in the isolated perfused rat heart. Marked differences in cardiac glycogen were produced by comparing hearts from rats previously treated with reserpine with hearts from control rats. Lesser differences in cardiac glycogen were produced in hearts by perfusing them for 15 minutes without glucose (0 mM glucose) or with 20 mM glucose. Both groups were then studied during a 5-minute anoxic cycle with 5 mM glucose as the exogenous substrate. Hearts from the reserpine-treated rats had higher left ventricular pressures, maximal rate of left ventricular pressure rise, and lactate output after 2 minutes of anoxia than the hearts from control rats. Similar but less marked mechanical differences were observed between 0 mM glucose and 20 mM glucose hearts. The mechanical differences during anoxia between the two groups were not abolished by simultaneous L-norepinephrine administration. Hearts with greater initial glycogen stores had higher glycogenolytic rates, and proportionately more lactate was produced from glycogen than from glucose. Thus, anaerobic ATP production per mole of hexose was greater in hearts with higher glycogen stores. Calculated ATP production was also greater in hearts from the reserpine-treated rats than in those from control animals. These studies demonstrate that both marked and minor elevations in cardiac glycogen are associated with greater glycolytic reserve and improved mechanical resistance to anoxia. This appears to be mainly due to enhanced glycogenolysis and anaerobic ATP production.

Isolation and determination of non-diffusible sialofucohexosaminoglycans derived from brain glycoproteins and their anatomical distribution in bovine brain.

Abstract— Glycoproteins in brain tissue were assayed by determining the amount of N‐acetylneuraminic acid (NANA), hexosamine, hexose, and fucose present in glycopeptides released by the proteolytic action of papain on the defatted protein residue that remains after treatment of the sample with chloroform‐methanol (2:1 and 1:2, v/v). Diffusible and non‐diffusible glycopeptides (sialofucohexosaminoglycans) were released by proteolysis. The procedure demonstrated that successive treatment of brain tissue with chloroform‐methanol (2:1, v/v) and chloroform‐methanol (1:2, v/v) removed all of the gangliosides present in the tissue.A 1 hr autolysis of rat brain tissue had no effect on the amount of glycopeptides recovered from the tissue. The carbohydrate composition of the non‐diffusible sialofucohexosaminoglycans was also unaffected.Areas of the brain that are enriched in neuronal cell bodies contained a higher concentration of gangliosides and glycoproteins than areas that consist largely of myelinated fibre tracts. On the other hand, there was a greater concentration of glycoprotein relative to that of gangliosides in areas that consist predominately of myelinated fibre tracts and glia than in areas enriched in neuronal cell bodies. The concentration of non‐diffusible sialofucohexosaminoglycans in whole bovine brain was less than that in whole rat brain. The non‐diffusible sialofucohexosaminoglycans from whole bovine brain contained less fucose and NANA per mole of hexosamine and hexose than non‐diffusible sialofucohexosaminoglycans isolated from whole rat brain. The non‐diffusible sialofucohexosaminoglycans isolated from bovine cerebral white matter were lower in fucose and NANA content per mole of hexose and hexosamine than those isolated from other brain areas. It is suggested that the fucose and NANA content of the non‐diffusible sialofucohexosaminoglycans associated with myelinated axons and (or) glia is less than that of the non‐diffusible sialofucohexosaminoglycans associated with the nerve cell body.

Composition, Properties, and Substrate Specificities of Myxobacter AL-1 Protease

Abstract Myxobacter strain AL-1 excretes a bacteriolytic enzyme designated as AL-1 protease. A procedure is described for obtaining homogeneous preparations of the enzyme. Physical studies revealed s020,w = 2.40 S; D020,w = 14 x 10-7 cm2 per sec; [n] = 1.66 ± 0.1 ml per g; a molecular weight of 14,300; frictional coefficient (f/f0) = 1.0. AL-1 protease contains 136 amino acids, a value which corresponds to a minimal molecular weight of 13,977. In contrast to many exocellular enzymes, this protein contains 1 disulfide bond, 1 mole of hexose, a relatively high content of aromatic amino acids, and an E1%1 cm = 15.8 at 280 mµ. The shortest peptide which served as a substrate was a tetrapeptide. Gly-Gly-Gly-Gly yielded Gly-Gly; Gly-Gly-Gly-Gly-Gly yielded Gly-Gly and Gly-Gly-Gly; l-Ala-Gly-Gly-Gly-Gly yielded l-Ala-Gly-Gly and Gly-Gly; Gly-Gly-Gly-Gly-l-Ala yielded Gly-Gly-Gly and Gly-l-Ala; Gly-Gly-l-Ala-Gly-Gly yielded Gly-Gly and l-Ala-Gly-Gly; and N-CBZ-Gly-l-Pro-l-Leu-Gly-l-Pro yielded N-CBZ-Gly-l-Pro and l-Leu-Gly-l-Pro. The hydrolysis points in the oxidized β chain of insulin were Ala-Leu, Val-CySO3H, and Gly-Phe.

Structural characteristics of the Fc′ fragment of human γG-globulin

The Fc′ fragment was isolated from normal human γG-globulin and a γG1(We) myeloma protein. It was shown to be heterogeneous by antigenic analysis, peptide mapping and C-terminal amino acid analysis. It had a molecular weight of 18,000 and contained 1·5 mole of hexose per mole of protein.