Crystal structure of the γ-cyclodextrin n-propanol inclusion complex; Correlation of α-, β-, γ-cyclodextrin geometries

Crystals of the hydrated n-propanol inclusion complex of γ-cyclodextrin (γ-CD; cyclo-octaamylose) have space group P4, a = b = 23.759(7), c = 23.069(7)Å and six quarter γ-CD per asymmetric unit. The structure was solved by YZARC and refined to R = 14% using 6300 X-ray counter data. The γ-CD are stacked, n-propanol (not located) occupies the channel-type cavity and 27 water sites populate interstices between stacks. Within the stacks γ-CD are arranged head-to-head as well as head-to-tail and H-bonded with O(2), O(3), O(6) hydroxyls. In the series α-,β-,γ-CD, angles C(1′)-O(4)-C(4) reduce from 119°-117.7°-112.6°, virtual O(4′)?O(4) distances increase 4.23-4.39-4.48 Å. intramolecular H-bonding distances O(2)?O(3) between adjacent glucoses, 3.00 Å in α-CD are wider than ～2.83 Å in β- and γ-CD, indicating a greater flexibility of the former.     

Effects of activation of μ-, κ<Subscript>1</Subscript>-, δ<Subscript>1</Subscript>-, δ<Subscript>2</Subscript>-, and ORL<Subscript>1</Subscript>-receptors on heart resistance to the pathogenic action of delayed ischemia and reperfusion

Different subtypes of opioid receptors (OR) were activated in rats in vivo to study the activation effect on the heart’s resistance to ischemia and reperfusion. It has been established that administration of deltorphin II, a selective δ₂-OR agonist, lowered the infarct size/area at risk index (IS/AAR) by 23%. Naltrexone, naloxone methiodide (an OR inhibitor not penetrating the blood-brain barrier (BBB)), and naltriben (δ₂-antagonist) eliminated the cardioprotective effect of deltorphin II, while BNTX (a δ₁-antagonist) produced no effect on the cardioprotective action of the δ₂-agonist. The infarct-reducing effect of deltorphin II was eliminated by administration of chelerythrine (a protein kinase C (PKC) inhibitor), glibenclamide (a K_ATP-channels inhibitor), and 5-hydroxydecanoate (a mitochondrial K_ATP-channel blocker). Administration of other opioids did not reduce the IS/AAR index. It has been established that all the deltorphins manifest antiarrhythmic potency. Other opioids do not produce any effect on the incidence of arrhythmia occurrences. The antiarrhythmic effect of deltorphin II was eliminated by preliminary administration of naltrexone, naloxone methiodide, and naltriben, but BNTX did not affect the δ₂-agonist’s anti-arrhythmic effect. The preliminary administration of chelerythrine, a PKC inhibitor, eliminated the δ₂ agonist’s antiarrhythmic action. However, glibenclamide and 5-hydroxydecanoate did not alter the antiarrhythmic effect by deltorphin II. Therefore, activation of the peripheral δ₂-ORs reduces the infarct size and prevents the onset of arrhythmias. The antiarrhythmic effect of the δ₂-OR stimulation is mediated by activating PKC and opening the mitochondrial K_ATP-channels. PKC participates in the antiarrhythmic effect of the δ₂-OR activation, but this effect does not depend on the condition of K_ATP-channels.     

α<Subscript>1</Subscript>-Thymosin, α<Subscript>2</Subscript>-interferon,and the LKEKK syntetic peptide inhibit the binding of the B subunit of the cholera toxin to intestinal epithelial cell membranes

The ¹²⁵I-labeled B-subunit of the cholera toxin ([125I]CT-B, specific activity of 98 Ci/mmol) was prepared. This subunit was shown to be bound to the membranes which were isolated from epithelial cells of a mucous tunic of the rat thin intestine with high affinity (K _d = 3.7 nM). The binding of the labeled protein was inhibited by the unlabeled α₂-interferon (IFN-α₂), α₁-thymosin, (TM-α₁), and the LKEKK synthetic peptide corresponding to the 16–20 sequence of TM-α₁ and the 131–135 sequence of human IFN-α₂ (Ki 1.0, 1.5, and 2.0 nM, respectively), whereas the KKEKL unlabeled synthetic peptide did not inhibit the binding (K _i > 100 μМ). The LKEKK peptide and CT-B were shown to dose-dependently increase an activity of the soluble guanylate cyclase (sGC) in the concentration range from 10 to 1000 nM. Thus, the binding of TM- α₁, IFN-α₂, and the LKEKK peptide to the CT-B receptor on a surface of the epithelial cells of the mucous tunic of the rat thin intestine resulted in an increase in the intracellular level of cGMP.     

Analyzing the Interaction of RseA and RseB, the Two Negative Regulators of the ��E Envelope Stress Response, Using a Combined Bioinformatic and Experimental Strategy

The Escherichia coli envelope stress response is controlled by the alternative sigma factor, σ^E, and is induced when unfolded outer membrane proteins accumulate in the periplasm. The response is initiated by sequential cleavage of the membrane-spanning antisigma factor, RseA. RseB is an important negative regulator of envelope stress response that exerts its negative effects onσ^E activity through its binding to RseA. In this study, we analyze the interaction between RseA and RseB. We found that tight binding of RseB to RseA required intact RseB. Using programs that performed global and local sequence alignment of RseB and RseA, we found regions of high similarity and performed alanine substitution mutagenesis to test the hypothesis that these regions were functionally important. This protocol is based on the hypothesis that functionally dependent regions of two proteins co-evolve and therefore are likely to be sequentially conserved. This procedure allowed us to identify both an N-terminal and C-terminal region in RseB important for binding to RseA. We extensively analyzed the C-terminal region, which aligns with a region of RseA coincident with the major RseB binding determinant in RseA. Both allele-specific suppression analysis and cysteine-mediated disulfide bond formation indicated that this C-terminal region of similarity of RseA and RseB identifies a contact site between the two proteins. We suggest a similar protocol can be successfully applied to pairs of non-homologous but functionally linked proteins to find specific regions of the protein sequences that are important for establishing functional linkage.The Escherichia coli σ^E-mediated envelope stress response is the major pathway to ensure homeostasis in the envelope compartment of the cell (1-3). σ^E regulon members encode periplasmic chaperones and proteases, the machinery for inserting β-barrel proteins into the outer membrane and components controlling the synthesis and assembly of LPS (4-6). This pathway is highly conserved among γ-proteobacteria (6).The σ^E response is initiated when periplasmic protein folding and assembly is compromised (7-9). During steady state growth, σ^E is inhibited by its antisigma factor, RseA, a membrane-spanning protein whose cytoplasmic domain binds to σ^E with picomolar affinity (10-13). Accumulation of unassembled porin monomers serves as a signal to activate the DegS protease to cleave RseA in its periplasmic domain (14, 15). This initiates a proteolytic cascade in which RseP cleaves periplasmically truncated RseA near or within the cytoplasmic membrane to release the RseA_cytoplasmic-σ^E complex, and cytoplasmic ATP-dependent proteases complete the degradation of RseA thereby releasing active σ^E (16-19).RseB, a second negative regulator of the envelope stress response (11, 20, 21), binds to the periplasmic domain of RseA with nanomolar affinity. RseB is an important regulator of the response (2, 22, 23). It prevents RseP from degrading intact RseA, thereby ensuring that proteolysis is initiated only when the DegS protease is activated by a stress signal (21). Additionally, RseB prevents activated DegS from cleaving RseA, suggesting that interaction of RseB with RseA must be altered before the signal transduction cascade is activated (23).The goal of the present studies was to explore how RseB binds to RseA. The interaction partner of RseB is the unstructured periplasmic domain of RseA (RseA-peri). Within RseA-peri, amino acids ∼169-186 constitute a major binding determinant to RseB (23, 24). This peptide alone binds RseB with 6 μm affinity, and deleting this region abrogates binding to RseB (23). Additional regions of RseA-peri also contribute to RseB binding, as intact RseA-peri binds with 20 nm affinity to RseB (23). Much less is known about the regions of RseB required for interaction with RseA. RseB is homodimeric two-domain protein, whose large N-terminal domain shares structural homology with LolA, a protein that transports lipoproteins to outer membrane (24, 25). The smaller C-terminal domain is connected to the N-terminal domain by a linker, and the two domains share a large interface, which may facilitate interdomain signaling. Glutaraldehyde cross-linking studies indicate that the C-terminal domain interacts with RseA, but the regions of interaction were not identified (25).In the present report, we study the interaction of RseB and RseA. We establish that both domains of RseB interact with RseA-peri. Using a global sequence alignment, we discovered several regions in RseA and RseB that had high sequence similarity, despite the low overall sequence similarity between these two proteins, a finding that was independently confirmed by a local sequence similarity algorithm. This suggested that these regions were functionally dependent, and we performed a set of mutagenesis experiments designed to test this idea. Our studies of the binding properties of these mutants revealed that regions in both the N terminus and C terminus of RseB modulate interaction with RseA. Moreover, genetic suppression analysis and cysteine-mediated disulfide bond formation suggest that the region of RseA/B with highest similarity (RseA residues 165-191 (major binding determinant in RseA) and RseB residues 233-258) are interacting partners.     

A double blind,randomized, placebo controlled study of the efficacy and safety of 5-Loxin<Superscript>®</Superscript>for treatment of osteoarthritis of the knee

Introduction

5-Loxin^® is a novel Boswellia serrata extract enriched with 30% 3-O-acetyl-11-keto-beta-boswellic acid (AKBA), which exhibits potential anti-inflammatory properties by inhibiting the 5-lipoxygenase enzyme. A 90-day, double-blind, randomized, placebo-controlled study was conducted to evaluate the efficacy and safety of 5-Loxin^® in the treatment of osteoarthritis (OA) of the knee.

Methods

Seventy-five OA patients were included in the study. The patients received either 100 mg (n = 25) or 250 mg (n = 25) of 5-Loxin^® daily or a placebo (n = 25) for 90 days. Each patient was evaluated for pain and physical functions by using the standard tools (visual analog scale, Lequesne''s Functional Index, and Western Ontario and McMaster Universities Osteoarthritis Index) at the baseline (day 0), and at days 7, 30, 60 and 90. Additionally, the cartilage degrading enzyme matrix metalloproteinase-3 was also evaluated in synovial fluid from OA patients. Measurement of a battery of biochemical parameters in serum and haematological parameters, and urine analysis were performed to evaluate the safety of 5-Loxin^® in OA patients.

Results

Seventy patients completed the study. At the end of the study, both doses of 5-Loxin^® conferred clinically and statistically significant improvements in pain scores and physical function scores in OA patients. Interestingly, significant improvements in pain score and functional ability were recorded in the treatment group supplemented with 250 mg 5-Loxin^® as early as 7 days after the start of treatment. Corroborating the improvements in pain scores in treatment groups, we also noted significant reduction in synovial fluid matrix metalloproteinase-3. In comparison with placebo, the safety parameters were almost unchanged in the treatment groups.

Conclusion

5-Loxin^® reduces pain and improves physical functioning significantly in OA patients; and it is safe for human consumption. 5-Loxin^® may exert its beneficial effects by controlling inflammatory responses through reducing proinflammatory modulators, and it may improve joint health by reducing the enzymatic degradation of cartilage in OA patients.

Trail Registration

(Clinical trial registration number: ISRCTN05212803.)     

SDF2L1, a Component of the Endoplasmic Reticulum Chaperone Complex, Differentially Interacts with ��-, ��-, and ��-Defensin Propeptides

Mammalian defensins are cationic antimicrobial peptides that play a central role in host innate immunity and as regulators of acquired immunity. In animals, three structural defensin subfamilies, designated as α, β, and θ, have been characterized, each possessing a distinctive tridisulfide motif. Mature α- and β-defensins are produced by simple proteolytic processing of their prepropeptide precursors. In contrast, the macrocyclic θ-defensins are formed by the head-to-tail splicing of nonapeptides excised from a pair of prepropeptide precursors. Thus, elucidation of the θ-defensin biosynthetic pathway provides an opportunity to identify novel factors involved in this unique process. We incorporated the θ-defensin precursor, proRTD1a, into a bait construct for a yeast two-hybrid screen that identified rhesus macaque stromal cell-derived factor 2-like protein 1 (SDF2L1), as an interactor. SDF2L1 is a component of the endoplasmic reticulum (ER) chaperone complex, which we found to also interact with α- and β-defensins. However, analysis of the SDF2L1 domain requirements for binding of representative α-, β-, and θ-defensins revealed that α- and β-defensins bind SDF2L1 similarly, but differently from the interactions that mediate binding of SDF2L1 to pro-θ-defensins. Thus, SDF2L1 is a factor involved in processing and/or sorting of all three defensin subfamilies.Mammalian defensins are tridisulfide-containing antimicrobial peptides that contribute to innate immunity in all species studied to date. Defensins are comprised of three structural subfamilies: the α-, β-, and θ-defensins (1). α- and β-Defensins are peptides of about 29–45-amino acid residues with similar three-dimensional structures. Despite their similar tertiary conformations, the disulfide motifs of α- and β-defensins differ. Expression of human α-defensins is tissue-specific. Four myeloid α-defensins (HNP1–4) are expressed predominantly by neutrophils and monocytes wherein they are packaged in granules, while two enteric α-defensins (HD-5 and HD-6) are expressed at high levels in Paneth cells of the small intestine. Myeloid α-defensins constitute about 5% of the protein mass of human neutrophils. HNPs are discharged into the phagosome during phagocytic ingestion of microbial particles. HD-5 and HD-6 are produced and stored as propeptides in Paneth cell granules and are processed extracellularly by intestinal trypsin (2). β-Defensins are produced primarily by various epithelia (e.g. skin, urogenital tract, airway) and are secreted by the producing cells in their mature forms. In contrast to pro-α-defensins, which contain a conserved prosegment of ∼40 amino acids, the prosegments in β-defensins vary in length and sequence. θ-Defensins are found only in Old World monkeys and orangutans and are the only known circular peptides in animals. These 18-residue macrocyclic peptides are formed by ligation of two nonamer sequences excised from two precursor polypeptides, which are truncated versions of ancestral α-defensins. Like myeloid α-defensins, θ-defensins are stored primarily in neutrophil and monocyte granules (3).Numerous laboratories have demonstrated that the antimicrobial properties of defensins derive from their ability to bind and disrupt target cell membranes (4), and studies have shown defensins to be active against Gram-positive and -negative bacteria (5), viruses (6–9), fungi (10, 11), and parasites such as Giardia lamblia (12). Defensins also play a regulatory role in acquired immunity as they are known to chemoattract T lymphocytes, monocytes, and immature dendritic cells (13, 14), act as adjuvants, stimulate B cell responses, and up-regulate proliferation and cytokine production by spleen cells and T helper cells (15, 16).Defensins are produced as pre-propeptides and undergo post-translational processing to form the mature peptides. While much has been learned about regulation of defensin expression, little is known about the factors involved in their biosynthesis. Valore and Ganz (17) investigated the processing of defensins in cultured cells and demonstrated that maturation of HNPs occurs through two proteolytic steps that lead to formation of mature α-defensins, but the proteases involved have yet to be identified. Moreover, there are virtually no published data regarding endoplasmic reticulum (ER)² factors that are responsible for the folding, processing, and sorting steps necessary for defensin maturation and secretion or trafficking to the proper subcellular compartment. It is likely that several chaperones, proteases, and protein-disulfide isomerase (PDI) family proteins are involved. Consistent with this possibility, Gruber et al. (18) recently demonstrated the role of a PDI in biosynthesis of cyclotides, small ∼30-residue macrocyclic peptides produced by plants.The primary structures of α- and θ-defensin precursors are closely related. We therefore undertook studies to identify proteins that interact with representative propeptides of each defensin subfamily with the goal of determining common and unique processes that regulate biosynthesis of α- and θ-defensins. We used two-hybrid analysis to first identify interactors of the θ-defensin precursor, proRTD1a. As described, we identified SDF2L1, a component of the ER-chaperone complex as an interactor, and showed that it also specifically interacts with α- and β-defensins. This suggests that SDF2L1 is involved in the maturation/trafficking of defensins at a step common to all three subfamilies of mammalian defensins.     

Analysis of the amide <Superscript>15</Superscript>N chemical shift tensor of the C<Subscript>α</Subscript> tetrasubstituted constituent of membrane-active peptaibols,the α-aminoisobutyric acid residue,compared to those of di- and tri-substituted proteinogenic amino acid residues

In protein NMR spectroscopy the chemical shift provides important information for the assignment of residues and a first structural evaluation of dihedral angles. Furthermore, angular restraints are obtained from oriented samples by solution and solid-state NMR spectroscopic approaches. Whereas the anisotropy of chemical shifts, quadrupolar couplings and dipolar interactions have been used to determine the structure, dynamics and topology of oriented membrane polypeptides using solid-state NMR spectroscopy similar concepts have been introduced to solution NMR through the measurements of residual dipolar couplings. The analysis of ¹⁵N chemical shift spectra depends on the accuracy of the chemical shift tensors. When investigating alamethicin and other peptaibols, i.e. polypeptides rich in α-aminoisobutyric acid (Aib), the ¹⁵N chemical shift tensor of this C_α-tetrasubstituted amino acid exhibits pronounced differences when compared to glycine, alanine and other proteinogenic residues. Here we present an experimental investigation on the ¹⁵N amide Aib tensor of N-acetyl-Aib-OH and for the Aib residues within peptaibols. Furthermore, a statistical analysis of the tensors published for di- (glycine) and tri-substituted residues has been performed, where for the first time the published data sets are compiled using a common reference. The size of the isotropic chemical shift and main tensor elements follows the order di- < tri- < tetra-substituted amino acids. A ¹⁵N chemical shift-¹H-¹⁵N dipolar coupling correlation NMR spectrum of alamethicin is used to evaluate the consequences of variations in the main tensor elements for the structural analysis of this membrane peptide.     

Integral role of the I′-helix in the function of the “inactive” C-terminal domain of catalase–peroxidase (KatG)

Catalase–peroxidases (KatGs) have two peroxidase-like domains. The N-terminal domain contains the heme-dependent, bifunctional active site. Though the C-terminal domain lacks the ability to bind heme or directly catalyze any reaction, it has been proposed to serve as a platform to direct the folding of the N-terminal domain. Toward such a purpose, its I′-helix is highly conserved and appears at the interface between the two domains. Single and multiple substitution variants targeting highly conserved residues of the I′-helix were generated for intact KatG as well as the stand-alone C-terminal domain (KatG^C). Single variants of intact KatG produced only subtle variations in spectroscopic and catalytic properties of the enzyme. However, the double and quadruple variants showed substantial increases in hexa-coordinate low-spin heme and diminished enzyme activity, similar to that observed for the N-terminal domain on its own (KatG^N). The analogous variants of KatG^C showed a much more profound loss of function as evaluated by their ability to return KatG^N to its active conformation. All of the single variants showed a substantial decrease in the rate and extent of KatG^N reactivation, but with two substitutions, KatG^C completely lost its capacity for the reactivation of KatG^N. These results suggest that the I′-helix is central to direct structural adjustments in the adjacent N-terminal domain and supports the hypothesis that the C-terminal domain serves as a platform to direct N-terminal domain conformation and bifunctionality.     

Biology of the electric eel, <Emphasis Type="Italic">Electrophorus electricus</Emphasis>, Linnaeus, 1766 (Gymnotiformes: Gymnotidae) on the floodplain of the Curiaú River,eastern Amazonia

The evaluation of the growth patterns and reproductive strategies of fish species are vitally important for the understanding of their biology and the management of stocks. The present study focused on the Amazonian electric eel (Electrophorus electricus), which is capable of producing an electrical discharge of up to 800 V. Specimens were collected on a monthly schedule from a floodplain in the eastern Amazon basin. The gonads of these specimens were examined, and the sex ratio, growth parameters, population structure, body size at first gonadal maturation, and the gonadosomatic index were determined. A balanced sex ratio was found. Males were larger than females, and both sexes presented isometric growth, which is unusual in species with an elongated anatomy. This isometry may be related to the reduction of the coelomic cavity, and its position near the head. The spawning period coincided with the start of the rainy season, and continued until high water, with the variation in gonadal development following the fluctuations in precipitation and river water levels. The asymptotic body length in both sexes was relatively large, and was inversely related to the growth coefficient (k), with a slower growth rate being recorded in the males. Mortality rates were relatively low in comparison with most species of tropical fish. The larger size of the male may be related to their role in parental care, and sexual selection on the part of the females. These findings may be important for the management of wild stocks, as well as captive rearing.     

Description of the male of Leptotyphlus kovaci ?ustek, 2000, the only Central European species of the Mediterranean genus Leptotyphlus Fauvel, 1874 (Coleoptera,Staphylinidae, Leptotyphlinae)

The previously unknown male of Leptotyphlus kovaci Šustek, 2000 is described and illustrated. The relationship of the species is discussed. The species is also reported from Gemerskoteplická jaskyňa near Jelšavská Teplica (Slovakia).     

Assessing the Efficiency of the Ability to Intentionally Variate the Power of β<Subscript>2</Subscript> Frequencies in the Frontal Lobes of the Cerebral Cortex

We performed longitudinal examinations by neurofeedback in 17 subjects. The subjects were trained for 12 training seßsions (three weeks) to voluntarily increase the intensity of the ß₂ frequencies in the frontal EEG electrodes of the right (the D scenario) and the left (the S scenario) hemispheres. All the subjects were divided into three groups depending on the training efficacy: a group of subjects that successfully controlled the ß activity in the frontal electrodes of both hemispheres (nine subjects), a group of subjects that successfully controlled this activity only in the right hemisphere (four subjects), and a group of subjects that failed to train during the specified period (four subjects). Analysis of the obtained data showed that the training efficacy depended on the cognitive activity that was focused on achieving the corresponding EEG effects and on the individual personality characteristics.     

Review of the fauna of Rotatoria,Cladocera, and Copepoda of the basin of the Anadyr’ River

The zooplankton composition is studied in the thermokarst, glacial and meteorite lakes, channels, former riverbeds, and hollows in the basin of Anadyr’. We found 174 taxa: 78, Rotatoria, 55, Cladocera, and 41, Copepoda. The most diverse is the lake fauna: 51 taxa of Rotatoria, 48, Cladocera, and 37, Copepoda. The thermokarst Lake Maiorskoe hosts 68 taxa: 31, Rotatoria, 14, Cladocera, and 23, Copepoda, wheras the cold ultraoligotrophic Lake El’gygytgyn features only one species of Cyclop of the group scutifer Cyclops neymanae Strel., and Rotatoria and Cladocera are present as allochtonous forms. The Copepoda illustrate the relations of the Anadyr’ fauna with those of Europe, North America, and Japan.     

Protease-catalyzed synthesis of the tripeptide CCK<Subscript>26–28</Subscript>, a fragment of CCK-8

Summary. Two enzymatically synthetic strategies of the tripeptide derivative PhAc-Asp(OMe)-Tyr-Met-OAl are reported. The second strategy gains the advantage of more economical starting materials, less reaction steps and a higher overall isolated yield of this tripeptide fragment over the first strategy. The effect of the acyl-donor ester concentration and structure, the C-α protecting group of the nucleophile, reaction media, enzyme and the carrier on the tripeptide derivative synthesis were studied. This tripeptide selected is a fragment of the cholecystokinin C-terminal octapeptide (CCK-8), a potential therapeutic agent in the control of gastrointestinal function and also a drug candidate for the treatment of epilepsy.     

Discovery of β-1,4-d-Mannosyl-N-acetyl-d-glucosamine Phosphorylase Involved in the Metabolism of N-Glycans

A gene cluster involved in N-glycan metabolism was identified in the genome of Bacteroides thetaiotaomicron VPI-5482. This gene cluster encodes a major facilitator superfamily transporter, a starch utilization system-like transporter consisting of a TonB-dependent oligosaccharide transporter and an outer membrane lipoprotein, four glycoside hydrolases (α-mannosidase, β-N-acetylhexosaminidase, exo-α-sialidase, and endo-β-N-acetylglucosaminidase), and a phosphorylase (BT1033) with unknown function. It was demonstrated that BT1033 catalyzed the reversible phosphorolysis of β-1,4-d-mannosyl-N-acetyl-d-glucosamine in a typical sequential Bi Bi mechanism. These results indicate that BT1033 plays a crucial role as a key enzyme in the N-glycan catabolism where β-1,4-d-mannosyl-N-acetyl-d-glucosamine is liberated from N-glycans by sequential glycoside hydrolase-catalyzed reactions, transported into the cell, and intracellularly converted into α-d-mannose 1-phosphate and N-acetyl-d-glucosamine. In addition, intestinal anaerobic bacteria such as Bacteroides fragilis, Bacteroides helcogenes, Bacteroides salanitronis, Bacteroides vulgatus, Prevotella denticola, Prevotella dentalis, Prevotella melaninogenica, Parabacteroides distasonis, and Alistipes finegoldii were also suggested to possess the similar metabolic pathway for N-glycans. A notable feature of the new metabolic pathway for N-glycans is the more efficient use of ATP-stored energy, in comparison with the conventional pathway where β-mannosidase and ATP-dependent hexokinase participate, because it is possible to directly phosphorylate the d-mannose residue of β-1,4-d-mannosyl-N-acetyl-d-glucosamine to enter glycolysis. This is the first report of a metabolic pathway for N-glycans that includes a phosphorylase. We propose 4-O-β-d-mannopyranosyl-N-acetyl-d-glucosamine:phosphate α-d-mannosyltransferase as the systematic name and β-1,4-d-mannosyl-N-acetyl-d-glucosamine phosphorylase as the short name for BT1033.     

Distribution of α-N-acetylgalactosaminidases among marine bacteria of the phylum <Emphasis Type="Italic">Bacteroidetes</Emphasis>, epiphytes of marine algae of the Seas of Okhotsk and Japan

Occurrence of α-N-acetylgalactosaminidases among 177 strains of marine bacteria of the phylum Bacteroidetes, epiphytes of marine algae growing on the littoral of the Seas of Okhotsk and Japan, was studied. About 36% of the isolates studied contained α-N-acetylgalactosaminidase. All of the bacteria of the genus Arenibacter (species A. latericius, A. certesii, and A. palladensis), irrespective of the source of isolation, synthesized this enzyme. The greatest number of α-N-acetylgalactosaminidase producers was found among the isolates from the algae Neosiphonia japonica, Acrosiphonia sonderi, and Ulva fenestrata sampled in the Cove of Trinity, Posyet Bay, the Sea of Japan. These were mainly bacteria of the genera Zobellia (50%) and Maribacter (58%). Among the epibionts studied, the bacteria Arenibacter latericius KMM 3523, an epiphyte of the brown alga Chorda filum from the Sea of Okhotsk, and Cellulophaga sp. KMM 6488, an epiphyte of the green alga Acrosiphonia sonderi from the Sea of Japan, were marked as the most promising sources of the enzyme. The results of this study showed that aerobic nonpathogenic marine Bacteroidetes, algal associants not requiring special cultivation conditions, are the promising, economical, and ecologically pure sources of unique and biotechnologically significant α-N-acetylgalactosaminidases.