Unbranched Polysaccharide Research Articles

Morphological changes induced by class III chitin synthase gene silencing could enhance penicillin production of Penicillium chrysogenum

Chitin synthases catalyze the formation of β-(1,4)-glycosidic bonds between N-acetylglucosamine residues to form the unbranched polysaccharide chitin, which is the major component of cell walls in most filamentous fungi. Several studies have shown that chitin synthases are structurally and functionally divergent and play crucial roles in the growth and morphogenesis of the genus Aspergillus although little research on this topic has been done in Penicillium chrysogenum. We used BLAST to find the genes encoding chitin synthases in P. chrysogenum related to chitin synthase genes in Aspergillus nidulans. Three homologous sequences coding for a class III chitin synthase CHS4 and two hypothetical proteins in P. chrysogenum were found. The gene which product showed the highest identity and encoded the class III chitin synthase CHS4 was studied in detail. To investigate the role of CHS4 in P. chrysogenum morphogenesis, we developed an RNA interference system to silence the class III chitin synthase gene chs4. After transformation, mutants exhibited a slow growth rate and shorter and more branched hyphae, which were distinct from those of the original strain. The results also showed that the conidiation efficiency of all transformants was reduced sharply and indicated that chs4 is essential in conidia development. The morphologies of all transformants and the original strain in penicillin production were investigated by light microscopy, which showed that changes in chs4 expression led to a completely different morphology during fermentation and eventually caused distinct penicillin yields, especially in the transformants PcRNAi1-17 and PcRNAi2-1 where penicillin production rose by 27 % and 41 %, respectively.

Pheromones: Fish Fear Factor

Fish, like many other animals, panic when another individual is injured. Now, the chemical nature of a substance that mediates this reaction has been uncovered.

Molecular structure of hyaluronan: an introduction

Hyaluronan is an unbranched polysaccharide of repeating disaccharides consisting of d-glucuronic acid and N-acetyl-d-glucosamine. Its strong water-retaining ability and visco-elastic properties have been broadly utilized in medical applications. Hyaluronan is an important constituent of the extracellular matrix whose physiological functions are manifested both as the substance is by itself as well as when it is being linked to various proteins. Compared with other biopolymers, such as nucleic acids and proteins, the structural chemistry of hyaluronan is much less developed. The scarce information about the metrical aspects of its structure shows no unusual features. Its secondary structure is characterized by intramolecular hydrogen bonding that is hard to distinguish from hydrogen bonding involving water molecules when hyaluronan is in aqueous medium. The tertiary structure of hyaluronan is sensitively dependent on its environment. The relative rigidity of the glycosidic bond and the intramolecular hydrogen bonds would tend to restrict rotational freedom and thus conformational variability. This, however, seems to be overwritten by the impact of molecular environment leading to a great variability of tertiary structure. A large number of conformations are possible and may be present as witnessed by their rather small free energy differences. Of the plethora of physical techniques and computational methods, X-ray crystallography and molecular dynamics calculations have proved to be the most fruitful so far. There are untapped possibilities in NMR spectroscopy for structural studies and quantum chemical calculations are also expected to contribute substantially to the structural chemistry of hyaluronan. There are many basic data as well as structural intricacies of hyaluronan that have so far eluded the researchers of its molecular structure.

The promise of chitosan microspheres in drug delivery systems

Chitosan is a partially deacetylated polymer obtained from the alkaline deacetylation of chitin, which is a glucose-based, unbranched polysaccharide that occurs widely in nature as the principal component of exoskeletons of crustaceans and insects, as well as of the cell walls of some bacteria and fungi. Chitosan exhibits a variety of physicochemical and biological properties resulting in numerous applications in fields such as waste water treatment, agriculture, fabric and textiles, cosmetics, nutritional enhancement and food processing. In addition to its lack of toxicity and allergenicity, its biocompatibility, biodegradability and bioactivity make it a very attractive substance for diverse applications as a biomaterial in the pharmaceutical and medical fields. This review takes a closer look at the biomedical applications of chitosan microspheres. Based on recent research and existing products, some new and potential future approaches in this fascinating area are discussed.

Therapeutic value of glycosaminoglycans in cancer

Glycosaminoglycans are unbranched polysaccharides composed of repeating units of alternating uronic acids and amino sugars. Most glycosaminoglycans are covalently attached to core proteins to form proteoglycans. Posttranslational modifications result in specific motifs that bind to a large variety of ligands, thus regulating growth factor signaling, cellular behavior, inflammation, angiogenesis, and the proteolytic environment. Dysregulated expression of glycosaminoglycans is present in cancer and reported to correlate with clinical prognosis in several malignant neoplasms. Recent knowledge on the biological roles of these molecules in cancer biology, tumor angiogenesis, and metastasis has promoted the development of drugs targeting them. Pharmaceutical approaches include the use of chemically modified heparins and glycosaminoglycans with defined structures, combination of inhibitors of glycosaminoglycan biosynthesis and polyamine depletion, and biologically active glycosaminoglycan-binding peptides. In addition, glycosaminoglycans are used as tumor-specific delivery and targeting vehicles for toxins and chemotherapeutics. Encouraging results in animal studies and clinical trials show the clinical relevance of glycosaminoglycan-based drugs and the use of glycosaminoglycans as therapeutic targets.

Potential applications of chitosan in veterinary medicine

Chitosan is a partially deacetylated polymer obtained from the alkaline deacetylation of chitin which is a glucose-based unbranched polysaccharide widely distributed in nature as the principal component of exoskeletons of crustaceans and insects as well as of cell walls of some bacteria and fungi. Chitosan exhibits a variety of physicochemical and biological properties resulting in numerous applications in fields such as waste and water treatment, agriculture, fabric and textiles, cosmetics, nutritional enhancement, and food processing. In addition to its lack of toxicity and allergenicity, and its biocompatibility, biodegradability and bioactivity make it a very attractive substance for diverse applications as a biomaterial in pharmaceutical and medical fields, where it has been used for systemic and local delivery of drugs and vaccines. It also has bioactive properties in its own right. This paper reviews current veterinary applications for chitosan including wound healing, bone regeneration, analgesic and antimicrobial effects. It also discusses the potential application of chitosan to drug and vaccine delivery in veterinary species. Given the restrictions imposed by financial and animal restraint considerations, especially in farming applications, the veterinary drug delivery areas most likely to benefit from chitosan are the delivery of chemotherapeutics such as antibiotics, antiparasitics, anaesthetics, painkillers and growth promotants to mucosal epithelium for absorption for local or systemic activity, and the delivery of immunomodulatory agents to the mucosal associated lymphoid tissue for induction or modulation of local immune responses. The properties of chitosan expected to enhance these functions are discussed, and the future research directions in this field are indicated.

NMR properties of alginate microbeads

Alginates are a family of unbranched polysaccharides with properties that vary widely depending on their composition. In the presence of multivalent cations (frequently Ca 2+), alginates form a gel. Consequently, alginates have been used to encapsulate a variety of biological materials, including cells. In this study, we present NMR relaxation and diffusion data from alginate microbeads with similar size and properties to those used in the development of a bioartificial pancreas. Our data demonstrate that the transverse relaxation time ( T 2) of water within the gel depends on the guluronic acid content of the alginate, whereas the longitudinal relaxation time ( T 1) and the apparent diffusion coefficient of water do not. Our data further suggest that the diffusion of Ca 2+ ions is hindered by the presence of a poly- l-lysine layer, a layer commonly added to provide mechanical support to the beads and immunoprotection to the encapsulated cells in the event of implantation. The impact of these data on our understanding of the role of alginate gels in the development of a bioartificial pancreas is discussed.

Glycogen Branching Enzyme Deficiency in Quarter Horse Foals

Seven related Quarter Horse foals that died by 7 weeks of age were examined for glycogen branching enzyme (GBE) deficiency. Clinical signs varied from stillbirth, transient flexural limb deformities, seizures, and respiratory or cardiac failure to persistent recumbency. Leukopenia (5 of 5 foals) as well as high serum creatine kinase (CK; 5 of 5), aspartate transaminase (AST; 4 of 4), and gamma glutamyl transferase (GGT; 5 of 5) activities were present in most foals, and intermittent hypoglycemia was present in 2 foals. Gross postmortem lesions were minor, except for pulmonary edema in 2 foals. Muscle, heart, or liver samples from the foals contained abnormal periodic acid Schiff's (PAS)‐positive globular or crystalline intracellular inclusions in amounts proportional to the foal's age at death. Accumulation of an unbranched polysaccharide in tissues was suggested by a shift in the iodine absorption spectra of polysaccharide isolated from the liver and muscle of affected foals. Skeletal muscle total polysaccharide concentrations were reduced by 30%, but liver and cardiac muscle glycogen concentrations were normal. Several glycolytic enzyme activities were normal, whereas GBE activity was virtually absent in cardiac and skeletal muscle, as well as in liver and peripheral blood cells of affected foals. GBE activities in peripheral blood cells of dams of affected foals and several of their half‐siblings or full siblings were ∼50% of controls. GBE protein in liver determined by Western blot was markedly reduced to absent in affected foals, and in a half‐sibling of an affected foal, it was approximately one‐half the amount of normal controls. Pedigree analysis also supported an autosomal recessive mode of inheritance. The affected foals have at least 2,600 half‐siblings. Consequently, GBE deficiency may be a common cause of neonatal mortality in Quarter Horses that is obscured by the variety of clinical signs that resemble other equine neonatal diseases.

Are hyaluronan receptors involved in three-dimensional cell migration?

Hyaluronan (HA), an unbranched polysaccharide consisting of repeated glucuronic acid/N-acetylglucosamine disaccharide units, is ubiquitously present in the extracellular matrix of many tissues (for a more comprehensive review see: Fraser et al., 1997). Increased amounts of hyaluronan are produced by solid tumors and tumor-associated fibroblasts, and tumor-induced HA is correlated with poor prognosis. HA is well known to stimulate the migration of a large variety of cell types. Stimulation of cell migration by HA has been explained by different mechanisms. HA was shown to specifically bind to cell surface receptors, and inhibition of HA-receptor function was demonstrated to decrease cell migration and tumor growth. On the other hand, HA as a large hydrophilic molecule is also known to modulate the extracellular packing of collagen and fibrin, leading to increased fiber size and porosity of extracellular substrates. Hence a modified matrix architecture might similarly account for increased locomotion of cells. In this review, we attempted to summarize the available data on HA-induced cell migration, with particular emphasis on the role of HA receptors in three-dimensional cell migration. Although the HA receptor CD44 has been shown to mediate migration of cells over two-dimensional hyaluronan-coated surfaces in vitro, there is only little evidence that HA-binding to CD44 or other HA receptors has major impact on the locomotion of cells through three-dimensional matrices in vivo. We showed recently that the promigratory effect of HA in fibrin gels is largely due to HA-mediated modulation of fibrin polymerization. By increasing the porosity of fibrin gels, HA strongly accelerates cell migration. The porosity of matrices therefore appears as an important and probably underestimated determinant of cell migration and tumor spread.

Phase Separation of Plant Cell Wall Polysaccharides and Its Implications for Cell Wall Assembly

Concentrated binary mixtures of polymers in solution commonly exhibit immiscibility, resolving into two separate phases each of which is enriched in one polymer. The plant cell wall is a concentrated polymer assembly, and phase separation of the constituent polymers could make an important contribution to its structural organization and functional properties. However, to our knowledge, there have been no published reports of the phase behavior of cell wall polymers, and this phenomenon is not included in current cell wall models. We fractionated cell walls purified from the pericarp of unripe tomatoes (Lycopersicon esculentum) by extraction with cyclohexane diamine tetraacetic acid (CDTA), Na2CO3, and KOH and examined the behavior of concentrated mixtures. Several different combinations of fractions exhibited phase separation. Analysis of coexisting phases demonstrated the immiscibility of the esterified, relatively unbranched pectic polysaccharide extracted by CDTA and a highly branched, de-esterified pectic polysaccharide present in the 0.5 N KOH extract. Some evidence for phase separation of the CDTA extract and hemicellulosic polymers was also found. We believe that phase separation is likely to be a factor in the assembly of pectic polysaccharides in the cell wall and could, for example, provide the basis for explaining the formation of the middle lamella.

The intercellular adhesin involved in biofilm accumulation of Staphylococcus epidermidis is a linear beta-1,6-linked glucosaminoglycan: purification and structural analysis

The primary attachment to polymer surfaces followed by accumulation in multilayered cell clusters leads to biofilm production of Staphylococcus epidermidis, which is thought to contribute to virulence in biomaterial-related infections. We purified a specific polysaccharide antigen of biofilm-producing S. epidermidis 1457 and RP62A, which was recently shown to have a function in the accumulative phase of biofilm production by mediating intercellular adhesion (D. Mack, M. Nedelmann, A. Krokotsch, A. Schwarzkopf, J. Heesemann, and R. Laufs, Infect. Immun. 62:3244-3253, 1994). Following Sephadex G-200 gel filtration, this antigen was separated by Q-Sepharose chromatography into a major polysaccharide, polysaccharide I (> 80%), which did not bind to Q-Sepharose, and a minor polysaccharide, polysaccharide II (< 20%), which was moderately anionic. As shown by chemical analyses and nuclear magnetic resonance spectroscopy, polysaccharide I is a linear homoglycan of at least 130 beta-1,6-linked 2-deoxy-2-amino-D-glucopyranosyl residues. On average, 80 to 85% of them are N acetylated; the rest are non-N-acetylating and positively charged. Chain cleavage by deamination with HNO2 revealed a more or less random distribution of the non-N-acetylated glucosaminyl residues, with some prevalence of glucosaminyl-rich sequences. Cation-exchange chromatography separated molecular species whose content of non-N-acetylated glucosaminyl residues varied between 2 and 26%. Polysaccharide II is structurally related to polysaccharide I but has a lower content of non-N-acetylated D-glucosaminyl residues and contains phosphate and ester-linked succinate, rendering it anionic. Enzyme-linked immunosorbent assay inhibition with various monosaccharides revealed the beta-anomeric form and the acetylated amino group of the D-glucosaminyl residues as important for reactivity with the specific antiserum. The unbranched polysaccharide structure favors long-range contacts and interactions between polysaccharide strands and the cell wall and/or lectin-like proteins, leading to intercellular adhesion and biofilm accumulation. The structure of the polysaccharide is, so far, considered to be unique and, according to its function, is referred to as S. epidermidis polysaccharide intercellular adhesin (PIA).

Identification of Structural Features of Heparin Required for Inhibition of Herpes Simplex Virus Type 1 Binding

Binding of HSV-1 to cells is mediated by interactions of virion glycoproteins gC and/or gB with heparan sulfate (HS) glycosaminoglycans on cell surface proteoglycans. HS and the related glycosaminoglycan, heparin, comprise a family of heterogeneous carbohydrates composed of long, unbranched polysaccharides modified, for example, by sulfations and acetylations. To define the specific features of HS important for viral binding, we took advantage of the structural similarities between heparin and cell surface HS and compared the ability of chemically modified heparin compounds to inhibit the binding of viral particles to the cell surface and subsequent plaque formation. Because binding presumably involves multiple, complex interactions between both known heparin-binding glycoproteins, gC and gB, and cell surface HS, we compared the effects of modified heparin compounds on the binding and subsequent plaque formation of wild-type and gC-negative strains of HSV-1 and, in select cases, the binding of gB-negative virus to cells. We identified specific structural features of heparin essential for the inhibition of viral binding. For example, both N-sulfation and 6-O-sulfation must be important determinants since desulfation of heparin at these sites abolished or decreased the antiviral activity of heparin. Moreover, we found that the antiviral activity of heparin was independent of its anticoagulant activity. Carboxyl-reduced and 2-,3-O-desulfated heparin selectively inhibited binding of gC-positive viruses (wild-type or a gB-negative strain) to cells, but had little or no inhibitory effect on binding and subsequent plaque formation for a gC-deletion virus. These results suggest that gC and gB interact with different structural features of HS.

Pharmacologic and Clinical Aspects of Intraarticular Injection of Hyaluronate

Hyaluronate (HA) is linear unbranched polysaccharide consisting of repeating disaccharide units of (1-4)-D-glucuronic acid-beta-(1-3)-D-N-acetylglucosamine. Extensive research has been conducted on HA, a major component of connective tissue. Hyaluronate, molecular weight 80 x 10(4)d, is available for the intraarticular injective treatment of osteoarthrosis of the knee and periarthritis of the shoulder. Hyaluronate relieves pain and has metabolic effects on articular cartilage, synovial tissue, and synovial fluid. Hyaluronate is a safe and effective treatment for patients with osteoarthrosis of the knee and periarthritis of the shoulder.

Proteoglycans: structure and function

Proteoglycans (PGs) occur predominantly in extracellular matrices and cell surfaces. Thcy consist of one or many glycosaminoglycan chains linked to protein. The glycosaminoglycans which are characteristic o f extracellular spaces include chondroitin sulphate, keratan sulphate, heparan sulphate, dermatan sulphate and hyaluronic acid ( H A ) [ I ] . Thcy are unbranched polysaccharides and all, with the exception of hyaluronate, occur in covalent linkage to protein. Their acidic nature arises from the presence of carboxylate and/or sulphate groups. T h e PGs in the extracellular matrix of cartilage have bcen the subject of much investigation and a great amount of information has become available in recent years on their molecular structure. T h e main bulk o f the tissue volume ih made up of the extracellular matrix and this consists o f a highly concentrated solution of PGs embedded in a network of type I1 collagen fibres. They can be extracted from cartilage in 4 M-guanidinium chloride in the presence of protcinasc inhibitors and purified by associative and dissociative density gradient centrifugation 121. T h e major PGs which occur in cartilage are the large, aggregating chondroitin sulphate PG and the smaller dermatan/chondroitin sulphate PGs. T h e former exists as a polydisperse structure with M , 0.5 x 10-4 x 10 D a 131. It consists of a protein core ( M , approx. 220 kDa) which is heavily substituted with chondroitin sulphate and keratan sulphate and with Nand 0linked oligosaccharides. T h e glycosaminoglycans are concentrated in domains along the protein core (Fig. I n ) . T h e keratan sulphate chains occupy a region close to the N terminus 141 and the chondroitin sulphate chains are found along a large portion of the polypeptide backbone stretching towards the C-terminus 151. T h e ability of this proteoglycan to form a strong and specific non-covalent interaction with H A (Fig. 1 h ) has been well characterized and has bcen shown to involve a globular domain at the N-terminus of the protein core [6, 71. A large number of proteoglycan monomers can bind to a single chain of HA. Doege er ul. [7a] have proposed the name 'aggrccan' to describe this structure. The resulting aggregated structure. with its very high negative-charge density, attracts a large amount of water so that it may occupy volumes that arc 30-50 times its dry weight [8]. T h e globular H A binding region ( H A B R ) binds t o a dccasaccharide or larger fragment of H A [c)]. A large number o f proteoglycans bind to a single hyaluronate chain and cach interaction is stabilized by link protein. T h e electrostatic repulsion between chondroitin chains on adjacent protco-

A computer-assisted structural analysis of regular polysaccharides on the basis of 13C-n.m.r. data

A computerised approach to the structural analysis of unbranched regular polysaccharides is described, which is based on an evaluation of the 13C-n.m.r. spectra for all possible primary structures within the additive scheme starting from the chemical shifts of the 13C resonances of the constituent monosaccharides and the average values of the glycosylation effects. The analysis reveals a structure (or structures), the evaluated spectrum of which resembles most closely that observed. The approach has been verified by using a series of bacterial polysaccharides of known structure and, in combination with methylation analysis data, for the determination of the presently unknown structures of the O-specific polysaccharides from Salmonella arizonae O59 and O63, and Proteus hauseri O19.

Structure of the O-chain polysaccharide of the phenol-phase soluble lipopolysaccharide of Escherichia coli 0:157:H7.

The phenol-phase soluble lipopolysaccharide isolated from Escherichia coli 0:157 by the hot phenol-water extraction procedure was shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, periodate oxidation, methylation, and 13C and 1H nuclear magnetic resonance studies to be an unbranched linear polysaccharide with a tetrasaccharide repeating unit having the structure: (formula; see text) The serological cross-reactivity of E. coli 0:157 with Brucella abortus, Yersinia enterocolitica (serotype 0:9), group N Salmonella, and some other E. coli species can be related immunochemically to the presence of 1,2-glycosylated N-acylated 4-amino-4, 6-dideoxy-alpha-D-mannopyranosyl residues in the O-chains of their respective lipopolysaccharides.

Proteoglycans: their structure, interactions and molecular organization in cartilage.

The term ‘proteoglycan’ was introduced in 1967 (see Balazs. 1970) to describe the family of molecules in which glycosaminoglycan chains are linked to protein. However. their rate of diffusion into textbooks of biochemistry has been rather slow, and I will therefore begin this presentation with a general introduction, which I hope will set the basis for their structural identity and indicate how their properties may be of importance in a broad biological context, before describing in more detail studies with proteoglycans from cartilage. Glycosaminoglycans are characteristic components of the extracellular space of vertebrate tissues. They are long unbranched polysaccharide chains which contain many acidic, carboxylate and/or sulphate groups. They do not normally occur as free chains in uiuo, but as proteoglycans, in which many like chains are linked by the reducing end of the terminal sugar residue to a protein. A whole technology was developed for the isolation, identification and quantification of glycosaminoglycan chains before their attachment to proteins was clearly established (Roden et al., 1972). Proteoglycan research thus began as a specialized area of carbohydrate chemistry. It is now apparent that in understanding the biological role of these macromolecules, the structure and interactions of the proteoglycans cannot be assessed by the analysis of the glycosaminoglycan chains alone. However, there are many difficulties in isolating and purifying intact proteoglycans, and it is only in recent years that a beginning has been made in exploring the full diversity of proteoglycan structure. There are seven types of glycosaminoglycan. on the basis of different repeating disaccharide units, which occur in mammalian tissues (Table I ) (see Muir & Hardingham, 1975). With the exception of hyaluronate, they have all been reported to occur linked to protein in different types of proteoglycan. The linkage to protein is via a neutral trisaccharide (Gal-Gal-Xyl) at the reducing end of the glycosaminoglycan chain, and xylose forms an 0-glycosidic bond with a serine residue in the protein. Keratan sulphate contains no neutral trisaccharide, but is linked via an 0-glycosidic bond between N-acetylgalactosamine to The Sixteenth Colworth Medal Lecture

Use of some new poly-phenolic resins for fractionation of carbohydrates and immobilisation of carbohydrate hydrolases and isomerases

New matrices, exhibiting selective adsorption of polymeric carbohydrates, have been synthesised by copolymerisation of hydroquinone and other phenolic compounds with formaldehyde or glutaraldehyde. These gel-type resins are capable of separating high molecular weight polysaccharides from low molecular weight oligosaccharides and monosaccharides, and branched from unbranched polysaccharides. The formaldehyde–hydroquinone resin is also an effective matrix for the simple and rapid immobilisation of various enzymes and (other) proteins. It is therefore additionally suitable for the separation of proteins from low molecular weight peptides and amino-acids. α-Amylase, glucoamylase, and D-glucose isomerase immobilised on the resins retain their activity incontinuous use. Since the resin matrices are highly porous and hydrophilic, they allow high flow rates to be achieved when used in packed-bed reactors. Both the formaldehyde and glutaraldehyde types of resin are cheap and easy to produce, relatively non-toxic in preparation and use, porous, hydrophilic, and extremely stable to changes in temperature and pH.

Structure of the Linkage Region between the Polysaccharide and Protein Parts of Saccharomyces cerevisiae Mannan

An oligosaccharide fragment corresponding to the linkage region between the polysaccharide and protein parts of Saccharomyces cerevisiae X2180 yeast mannan has been isolated. This was accomplished by the combined action of a bacterial endo-α(1→6)mannanase and an endo-β-N-acetylglucosaminidase on the mutant mannan from S. cerevisiae X2180-1A-5 that has an unbranched polysaccharide outer chain. The mannanase removed the unbranched mannan outer chain and the glucosaminidase apparently split a di-N-acetylchitobiose unit at the point of attachment of the polysaccharide to asparagine in the protein part. This order of degradation is assumed from the facts that wild type mannan with a branched outer chain was not attacked whereas ovalbumin was degraded with the release of the expected oligosaccharide fragment terminated by N-acetylglucosamine at the reducing end. Thus, the glucosaminidase was able to act only on a partially degraded mannan chain. The linkage fragment was composed of about 12 mannose units and a single N-acetylglucosamine which was found at the reducing end. The attachment of mannose to N-acetylglucosamine was by a β(1→4) linkage as in other glycoproteins. Acetolysis of the oligosaccharide yielded mannose, mannobiose αMan(1→2)Man and αMan(1→3)Man, mannotriose with both 1→2 and 1→3 linkages, and mannotetraose, probably αMan(1→3)αMan(1→2)αMan(1→3)Man. These fragments were interlinked by α(1→6) bonds, and the oligosaccharide was attached to the N-acetylglucosamine. Another acetolysis fragment isolated, αMan(1→3)αMan(1→2)αMan(1→3)βMan(1→4)GNAc, is common to other glycoproteins. The observation that this kind of branched inner core structure is made in a yeast mutant that forms an unbranched outer chain demonstrates that the formation of these two parts of the molecule is regulated in part by different enzymes and may serve different functions in the translocation of the mannan proteins and their organization in the cell wall.

Substrate specificity of mycodextranase.

MYCODEXTRAN (nigeran) is an unbranched polysaccharide composed of a regular alternating sequence of α–(1rarr;3) and α–(1→4) linked glucopyranosyl units. It is produced in the mycelia of various species of moulds1. Reese and Mandels have recently reported2 the isolation and partial purification of an inducible glucanase (mycodextranase) produced in the culture filtrates of selected species of fungi grown on a medium containing mycodextran as a carbon source. The enzyme is an endoglucanase that hydrolyses nigeran to produce hexameric and tetrameric units which are subsequently hydrolysed to produce the disaccharide nigerose (O–α–D–glucopyranosyl–(1→3)–D–glucose) as the final product2.