The Clostridium cellulovorans cellulosome: an enzyme complex with plant cell wall degrading activity

Cellulose comprises a major portion of biomass on the earth, and the turnover of this material contributes to the CO2 cycle. Cellulases, which play a major role in the turnover of cellulosic materials, have been found either as free enzymes that work synergistically, or as an enzyme complex called the cellulosome. This review summarizes some of the general properties of cellulosomes, and more specifically, the properties of the Clostridium cellulovorans cellulosome. The C cellulovorans cellulosome is an extracellular enzyme complex with a molecular weight of about 1 x 10(6), and is comprised of at least ten subunits. The major subunit is the scaffolding protein CbpA, with a molecular weight of 189,000. This nonenzymatic subunit contains a cellulose binding domain (CBD) that binds the cellulosome to the substrate, nine conserved cohesins or enzyme binding domains, and four conserved surface layer homologous (SLH) domains. It is postulated that the SLH domains help to bind the cellulosome to the cell surface. The cellulosomal enzymes include cellulases (family 5 and 9 endoglucanases and a family 48 exoglucanase), a mannanase, a xylanase, and a pectate lyase. The cellulosome is capable of converting Arabidopsis and tobacco plant cells to protoplasts. One of the endoglucanases, EngE, contains three tandemly repeated SLHs at its N-terminus, and therefore appears capable of binding to the scaffolding protein CbpA as well as to the cell surface. Cellulosomes can attack crystalline cellulose, but the free cellulosomal enzymes can attack only soluble and amorphous celluloses. Nine genes for the cellulosome are found in a gene cluster cbpA-exgS-engH-engK-hbpA-engL-manA-engM-engN. Other cellulosomal genes such as engB, engE, and engY are not linked to the major gene cluster or to each other. By determining the structure and function of the cellulosome, we hope to increase the efficiency of the cellulosome by genetic engineering techniques.     

N-Allyl-N-tert-butyldimethylsilylamine for chiral ligand-controlled asymmetric conjugate addition to tert-butyl alkenoates

The chiral ligand controlled asymmetric conjugate addition reaction of lithium N-allyl-N-(tert-butyldimethylsilyl)amide to alkenoates proceeded smoothly to give, after protodesilylation, the corresponding 3-allylaminoalkanoates with high enantioselectivities in high yields. The allyl group on the nitrogen atom was easily removable to afford 3-aminoalkanoates.     

Magnetic susceptibility and specific heat of uranium double perovskite oxides Ba2MUO6 (M=Co, Ni)

Double perovskites Ba₂MUO₆ (M=Co, Ni) were prepared by the solid-state reaction. X-ray diffraction measurements show that both cobalt (nickel) and uranium ions are ordered in the NaCl type over the six-coordinate B sites of the perovskite ABO₃. Detailed magnetic susceptibility and specific heat measurements show that Ba₂CoUO₆ and Ba₂NiUO₆ order ferromagnetically at 9.1 and 25 K, respectively. From the analysis of the magnetic specific heat, the ground states of the Co²⁺ and Ni²⁺ ions were determined.     

Spongelike Macroporous TiO2 Monoliths Prepared from Starch Gel Template

TiO₂ materials with a hierarchical meso/macropore organization were fabricated by using titanium dioxide nanoparticles and starch gel templates. Starch sponges with high internal macroporosities were prepared by freezing and thawing of starch gels and were then infiltrated with colloidal suspensions of titania nanoparticles and air-dried to produce TiO₂-starch foams with pores up to 200 m across depending on the starch concentration and the TiO₂ loading. The TiO₂ nanoparticles were deposited as coherent layers on the thin walls of the starch framework, which was removed by calcination without significant disruption of the TiO₂ framework. The three dimensional macroporous structures of TiO₂-starch sponge composite and TiO₂ sponge induced extremely high photocatalytic activity.     

Alternating polyampholytes prepared by hydrolysis of copolymer of maleic anhydride and N-vinylsuccinimide

Alternating polyampholytes (MA-VA) containing two acidic groups and one basic group were prepared by the copolymerization of maleic anhydride (M₁) and N-vinylsuccinimide (M₂) at 60°C with AIBN as the initiator, followed by acid hydrolysis with 1N hydrochloric acid at 140°C for 24 hr. The monomer reactivity ratios r₁ and r₂ are 0.025 and 0.06, respectively. The structure of polymers was discussed on the basis of the data of their elementary, infrared (IR), and thermal analyses and the binding ability of heavy metal ion. Polyampholytes were soluble in strong acidic and basic media but were precipitated in the pH range 3–4. An isoelectric point at pH 3 was determined by potentiometric titration and the turbidimetric method. By thermal treatment above 205°C the polyampholyte turned quantitatively into a cyclized lactam. This suggests that the polyampholyte MA–VA has an intramolecular hydrogen bond between the amino and γ-carboxyl groups. The binding of Cu²⁺ and Hg²⁺ by the polyelectrolyte was evaluated by equilibrium dialysis.     

Nickel(0)-mediated sequential addition of carbon dioxide and aryl aldehydes into terminal allenes

[reaction: see text] Nickel-mediated sequential addition of carbon dioxide and aryl aldehydes into terminal allenes is reported. The reaction proceeded in a diastereoselective manner to afford alpha-methylene-gamma-hydroxy carboxylic acids, which allowed stereoselective preparation of cis-beta, gamma-disubstituted alpha-methylene-gamma-lactones.     

Preparation of O-1∼C-6 and O-7∼C-14 fragments of colletodiol

(5S,2E)-5-Tetrahydropyranyloxy-2-hexenoic acid and p-toluenesulfonylethyl (4R,5R,7R,2E)-7-hydroxy-4,5-dimethylmethylenedioxy-2-octenoate were prepared from ethyl acetoacetate and D-glucose, respectively.     

Novel Mn(II)Mn(III)Mn(II) trinuclear complexes with carbohydrate bridges derived from seven-coordinate manganese(II) complexes with N-glycoside

Reactions of MnX2.nH2O with tris(N-(D-mannosyl)-2-aminoethyl)amine ((D-Man)3-tren), which was formed from D-mannose and tris(2-aminoethyl)amine (tren) in situ, afforded colorless crystals of [Mn((D-Man)3-tren)]X2 (3a, X = Cl; 3b, X = Br; 3c, X = NO3; 3d, X = 1/2SO4). The similar reaction of MnSO4.5H2O with tris(N-(L-rhamnosyl)-2-aminoethyl)amine ((L-Rha)3-tren) gave [Mn((L-Rha)3-tren)]SO4 (4d), where L-rhamnose is 6-deoxy-L-mannose. The structures of 3b and 4d were determined by X-ray crystallography to have a seven-coordinate Mn(II) center ligated by the N-glycoside ligand, (aldose)3-tren, with a C3 helical structure. Three D-mannosyl residues of 3b are arranged in a delta(ob3) configuration around the metal, leading to formation of a cage-type sugar domain in which a water molecule is trapped. In 4d, three L-rhamnosyl moieties are in a delta(lel3) configuration to form a facially opened sugar domain on which a sulfate anion is capping through hydrogen bonding. These structures demonstrated that a configurational switch around the seven-coordinate manganese(II) center occurs depending on its counteranion. Reactions of 3a, 3b, and 4d with 0.5 equiv of Mn(II) salt in the presence of triethylamine yielded reddish orange crystals formulated as [[Mn((aldose)3-tren)]2Mn(H2O)X3.nH2O (5a, aldose = D-Man, X = Cl; 5b, aldose = D-Man, X = Br; 6d, aldose = L-Rha, X = 1/2SO4). The analogous trinuclear complexes 6a (aldose = L-Rha, X = Cl), 6b (aldose = L-Rha, X = Br), and 6c (aldose = L-Rha, X = NO3) were prepared by the one-pot reaction of Mn(II) salts with (L-Rha)3-tren without isolation of the intermediate Mn(II) complexes. X-ray crystallographic studies revealed that 5a, 5b, 6c, and 6d have a linearly ordered trimanganese core, Mn(II)Mn(III)Mn(II), bridged by two carbohydrate residues with Mn-Mn separations of 3.845(2)-3.919(4) A and Mn-Mn-Mn angles of 170.7(1)-173.81(7) degrees. The terminal Mn(II) atoms are seven-coordinate with a distorted mono-face-capped octahedral geometry ligated by the (aldose)3-tren ligand through three oxygen atoms of C-2 hydroxyl groups, three N-glycosidic nitrogen atoms, and a tertiary amino group. The central Mn(III) atoms are five-coordinate ligated by four oxygen atoms of carbohydrate residues in the (aldose)3-tren ligands and one water molecule, resulting in a square-pyramidal geometry. In the bridging part, a beta-aldopyranosyl unit with a chair conformation bridges the two Mn(II)Mn(III) ions with the C-2 mu-alkoxo group and with the C-1 N-glycosidic amino and the C-3 alkoxo groups coordinating to each metal center. These structures could be very useful information in relation to xylose isomerases which promote aldose-ketose isomerization by using divalent dimetal centers such as Mn2+, Mg2+, and Co2+.     

Oxidative Degradation of trans‐1,4‐Polyisoprene Cast Films and Single Crystals by Enzyme‐Mediator Systems

Depolymerization and morphological changes of cast films and single crystals of trans‐1,4‐polyisoprene with three enzyme‐mediator systems, lipoxygenase/linoleic acid (LPO/LH), horseradish peroxidase/1‐hydroxybenzotriazole (HRP/1‐HBT) and laccase/1‐hydroxybenzotriazole/linoleic acid (laccase/1‐HBT/LH), were investigated by scanning electron microscopy and gel permeation chromatography. Treatment of cast film with a LPO/LH system involving radical generation via lipid peroxidation led to a marked decrease in the molecular weight of polymers. LPO/LH system resulted in the morphological damage of single crystals of trans‐1,4‐polyisoprene. Laccase/1‐HBT/LH also destroyed the single crystals and depolymerized emulsified trans‐1,4‐polyisoprene. On the contrary, spherulites appeared on the surface of cast films and the molecular weight slightly decreased after treatment with HRP/1‐HBT, indicating that the amorphous region was predominantly degraded and the crystal region remained unchanged. The morphology of single crystals remained unchanged during the treatment of HRP/1‐HBT system for 6 d. In addition, Fenton reagent/linoleic acid was used as a chemical initiator of lipid peroxidation for degradation of trans‐1,4‐polyisoprene cast films. This system reduced molecular weight of the cast films, as well as LPO/LH system.

Scanning electron micrographs of trans‐1,4‐polyisoprene single crystals before (A) and after (B) treatment with the lipoxygenase/linoleic acid system.