Nerve block efficacy in breast augmentation: A systematic review and meta-analysis.

Breast augmentation is often performed as a day-case general anaesthetic operation, with postoperative, opioid-based analgesia regimens. However, it may also be performed using regional anaesthesia; a variety of nerve block techniques are available to reduce postoperative pain and analgesic requirements. This systematic review and meta-analysis were undertaken according to the Preferred Reporting Items for Systematic Reviews and Meta-analysis guidelines comparing breast augmentation using regional anaesthesia with general anaesthesia, versus general anaesthesia alone or with local field infiltration. All randomised or quasi-randomised studies that recruited adult female patients undergoing breast augmentation using regional anaesthesia were considered. The primary outcome measures were postoperative pain and analgesic requirements. A randomised effects model was used, with standardised mean difference or mean difference outcomes used as appropriate. Thirteen studies were included for systematic review, out of which eight met the inclusion criteria for meta-analysis. Nerve blocks had statistically significant standardised mean difference reductions in postoperative pain scores across all time points: 0 h (-1.2 [-2.1 to -0.3], p = 0.01, I2 = 85%), 1 h (-1.3 [-2.1 to -0.5], p = 0.002, I2 = 89%), 2 h (-1.8 [-2.8 to -0.9], p = 0.0002, I2 = 88%), 4-6 h (-1.2 [-2.1 to -0.4], p = 0.006, I2 = 89%), 24 h (-1.4 [-2.5 to -0.2], p = 0.02, I2 = 94%). There was also a statistically significant reduction in postoperative opioid requirements: -150 mcg fentanyl (-259.2 to -40.9), p = 0.007. Although an element of study heterogeneity is noted, this systematic review and meta-analysis support the concept that regional anaesthesia using nerve blocks in breast augmentation surgery, reduces both postoperative pain and opioid requirements, compared with general anaesthesia.

Platelet-Rich Plasma (PRP) for Sacrococcygeal Pilonidal Disease: An Updated Systematic Review and Meta-Analysis.

BACKGROUND: Pilonidal disease can be a debilitating condition which carries a significant physical and economic burden. This systematic review and updated meta-analysis presents the evidence for the use of platelet-rich plasma (PRP) for wound healing following open and minimally-invasive sacrococcygeal pilonidal surgery. METHODS: A literature search was performed during December 2021 for studies relating to platelet-rich plasma and pilonidal wound healing following surgery. RESULTS: Nine studies remained after applying the exclusion criteria, incorporating a total of 621 (open surgery group) and 309 (minimally-invasive group) patients, respectively. Pooled analysis of the six open surgery group studies demonstrated a significant reduction in wound healing time (mean difference [MD] = - 13.98 days, 95% CI - 18.41 to - 9.55, p < 0.001, I2 = 98%). Three open surgery group studies compared post-operative time off work, while three recorded mean pain duration; pooled analysis also revealed a significant reduction in both outcomes, respectively (MD = - 8.7 days, 95% CI - 9.4 to - 8.0, p < 0.001, I2 = 57%; MD = - 9.5 days, 95% CI - 15.6 to - 3.3, p = 0.002, I2 = 98%). Methodological heterogeneity among the minimally-invasive studies precluded formal meta-analysis; however, two studies demonstrated a modest improvement in wound healing when treated with PRP. CONCLUSIONS: This systematic review and updated meta-analysis provide further evidence supporting the use of PRP for wound healing in sacrococcygeal pilonidal disease. PRP application was demonstrated to significantly reduce healing time, postoperative pain and time off work in the open surgery group. Nevertheless, there is still considerable heterogeneity among PRP manufacture and administration techniques, and further high-powered RCTs with consistent methodology are required to substantiate these findings.

Chyle leak following total colectomy for ulcerative colitis: a case report and review of the literature.

Chyle leak is a rare complication in colorectal surgery. It occurs due to disruption of the lymphatic drainage network in the abdomen or retroperitoneum. We describe the first reported case of chyle leak following total colectomy for inflammatory bowel disease. Our patient underwent total colectomy for severe ulcerative colitis not responsive to medical treatment. Four days postoperatively, a milky fluid was noted in the drainage bag. Analysis of the fluid confirmed chyle. The patient remained well and was successfully managed conservatively with a fat-free elemental diet and was discharged from hospital on day 12 postoperatively. A review of the literature suggests that conservative management with dietary modification is a common and effective management strategy; however, medical and surgical options exist for refractory cases.

Purple Urine Bag Syndrome in a Patient with an Ileal Conduit and Clostridium Difficile Infection.

Purple urine bag syndrome is a potentially alarming phenomenon caused by bacterial metabolism of urinary tryptophan into indigo (blue) and indirubin (red) pigments. We report the case of a 46-year-old female with an ileal conduit who presented with a 2 week history of abdominal pain and purple discolouration of her urine. In addition, we review the literature on purple urine bag syndrome, and identify potential new risk factors and management considerations.

Phosphorylated human lectin galectin-3: analysis of ligand binding by histochemical monitoring of normal/malignant squamous epithelia and by isothermal titration calorimetry.

The human lectin galectin-3 is a multifunctional effector with special functions in regulation of adhesion and apoptosis. Its unique trimodular organization includes the 12-residue N-terminal sequence, a substrate for protein kinase CK1-dependent phosphorylation. As a step towards elucidating its significance, we prepared phosphorylated galectin-3, labelled it and used it as a tool in histochemistry. We monitored normal and malignant squamous epithelia. Binding was suprabasal with obvious positive correlation to the degree of differentiation and negative correlation to proliferation. The staining pattern resembled that obtained with the unmodified lectin. Basal cell carcinomas were invariably negative. The epidermal positivity profile was akin to distribution of the desmosomal protein desmoglein, as also seen with keratinocytes in vitro. In all cases, binding was inhibitable by the presence of lactose, prompting further investigation of the activity of the lectin site by a sensitive biochemical method, i.e. isothermal titration calorimetry. The overall affinity and the individual enthalpic and entropic contributions were determined. No effect of phosphorylation was revealed. This strategic combination of histo- and biochemical techniques applied to an endogenous effector after its processing by a protein kinase thus enabled a detailed monitoring of the binding properties of the post-translationally modified lectin. It underscores the value of using endogenous lectins as a histochemical tool. The documented approach has merit for applications beyond lectinology.

Directing the immune response to carbohydrate antigens.

Peptide mimetics may substitute for carbohydrate antigens in vaccine design applications. At present, the structural and immunological aspects of antigenic mimicry, which translate into immunologic mimicry, as well as the functional correlates of each, are unknown. In contrast to screening peptide display libraries, we demonstrate the feasibility of a structure-assisted vaccine design approach to identify functional mimeotopes. By using concanavalin A (ConA), as a recognition template, peptide mimetics reactive with ConA were identified. Designed peptides were observed to compete with synthetic carbohydrate probes for ConA binding, as demonstrated by enzyme-linked immunosorbent assay and isothermal titration calorimetry (ITC) analysis. ITC measurements indicate that a multivalent form of one particular mimetic binds to ConA with similar affinity as does trimannoside. Splenocytes from mimeotope-immunized mice display a peptide-specific cellular response, confirming a T-cell-dependent nature for the mimetic. As ConA binds to the Envelope protein of the human immunodeficiency virus, type 1 (HIV-1), we observed that mimeotope-induced serum also binds to HIV-1-infected cells, as assessed by flow cytometry, and could neutralize T-cell line adapted HIV-1 isolates in vitro, albeit at low titers. These studies emphasize that mimicry is based more upon functional rather than structural determinants that regulate mimeotope-induced T-dependent antibody responses to polysaccharide and emphasize that rational approaches can be employed to develop further vaccine candidates.

Multivalent protein-carbohydrate interactions. A new paradigm for supermolecular assembly and signal transduction.

Many biological recognition processes involve the binding and clustering of ligand-receptor complexes and concomitant signal transduction events. Such interactions have recently been observed in human T cells in which binding and cross-linking of specific glycoprotein counter-receptors on the surface of the cells by an endogenous bivalent carbohydrate binding protein (galectin-1) leads to apoptosis [Pace, K. E., et al. (1999) J. Immunol. 163, 3801-3811]. Importantly, different counter-receptors associated with specific phosphatase or kinase activities were shown to form separate clusters on the surface of the cells as a result of galectin-1 binding to the carbohydrate moieties of the respective glycoproteins. This suggests that the unique separation and organization of signaling molecules that results from galectin-1 binding is involved in delivering the signal to die. The ability of galectin-1 to induce the separation of specific glycoprotein receptors was modeled on the basis of molecular and structural studies of the binding of multivalent carbohydrates to lectins that result in the formation of specific two- and three-dimensional cross-linked lattices. These latter studies have been recently highlighted by X-ray crystallographic results showing that a single tetravalent lectin forms distinct cross-linked complexes with four different bivalent oligosaccharides [Olsen, L. R., et al. (1997) Biochemistry 36, 15073-15080]. In this report, binding and cross-linking of multivalent carbohydrates with multivalent lectins is shown to be a new paradigm for supermolecular assembly and signal transduction in biological systems.

Lectin cross-linking interactions with multivalent carbohydrates.

The present findings provide a molecular basis for a new paradigm of specificity in multivalent carbohydrate-lectin interactions, namely the formation of type 2 homogeneous cross-linked lattices between multivalent carbohydrates and lectins. The present x-ray data demonstrate that the cross-linked complexes formed between a series of structurally related divalent carbohydrates and a single tetravalent lectin (SBA) are distinct and due to crystal packing interactions. These results thus provide a molecular basis for the formation of homogeneous type 2 cross-linked complexes between lectins and multivalent carbohydrates and glycoconjugates. These findings are also relevant to the observations that lectin-carbohydrate cross-linking interactions are involved in cellular recognition and signal transduction processes. For example, activated human T-cells undergo apoptosis due to binding and cross-linking of specific glycoprotein receptors by galectin-1 (Pace et al., 1999). Confocal microscopy shows that the galectin cross-linked glycoprotein receptors form homogeneous aggregates from a population of previously dispersed molecules on the surface of the cells. The crystal structures of the four SBA/pentasaccharide complexes thus repesent models for lectin-carbohydrate clustering in vivo.

Binding of multivalent carbohydrates to concanavalin A and Dioclea grandiflora lectin. Thermodynamic analysis of the "multivalency effect".

Binding of a series of synthetic multivalent carbohydrate analogs to the Man/Glc-specific lectins concanavalin A and Dioclea grandiflora lectin was investigated by isothermal titration microcalorimetry. Dimeric analogs possessing terminal alpha-D-mannopyranoside residues, and di-, tri-, and tetrameric analogs possessing terminal 3, 6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranoside residues, which is the core trimannoside of asparagine-linked carbohydrates, were selected in order to compare the effects of low and high affinity analogs, respectively. Experimental conditions were found that prevented precipitation of the carbohydrate-lectin cross-linked complexes during the isothermal titration microcalorimetry experiments. The results show that the value of n, the number of binding sites on each monomer of the lectins, is inversely proportional to the number of binding epitopes (valency) of each carbohydrate. Hence, n values close to 1.0, 0.50, and 0.25 were observed for the binding of mono-, di-, and tetravalent sugars, respectively, to the two lectins. Importantly, differences in the functional valency of a triantennary analog for concanavalin A and D. grandiflora lectin are observed. The enthalpy of binding, DeltaH, is observed to be directly proportional to the number of binding epitopes in the higher affinity analogs. For example, DeltaH of a tetravalent trimannoside analog is nearly four times greater than that of the corresponding monovalent analog. Increases in K(a) values of the multivalent carbohydrates relative to monovalent analogs, known as the "multivalency effect," are shown to be due to more positive entropy (TDeltaS) contributions to binding of the former sugars. A general thermodynamic model for distinguishing binding of multivalent ligands to a single receptor with multiple, equal subsites versus binding to separate receptor molecules is given.

Thermodynamic binding studies of lectins from the diocleinae subtribe to deoxy analogs of the core trimannoside of asparagine-linked oligosaccharides.

Lectins from seven different species of the Diocleinae subtribe have been recently isolated and characterized in terms of their carbohydrate binding specificities (Dam, T. K., Cavada, B. S., Grangeiro, T. B., Santos, C. F., de Sousa, F. A. M., Oscarson, S., and Brewer, C. F. (1998) J. Biol. Chem. 273, 12082-12088). The lectins included those from Canavalia brasiliensis, Cratylia floribunda, Dioclea rostrata, Dioclea virgata, Dioclea violacea, and Dioclea guianensis. All of the lectins exhibited specificity for Man and Glc residues, but much higher affinities for the branched chain trimannoside, 3,6-di-O-(alpha-d-mannopyranosyl)-d-mannose, which is found in the core region of all asparagine-linked carbohydrates. In the present study, isothermal titration microcalorimetry is used to determine the binding thermodynamics of the above lectins, including a new lectin from Canavalia grandiflora, to a complete series of monodeoxy analogs of the core trimannoside. From losses in the affinity constants and enthalpies of binding of certain deoxy analogs, assignments are made of the hydroxyl epitopes on the trimannoside that are involved in binding to the lectins. The pattern of binding of the deoxy analogs is similar for all seven lectins, and similar to that of concanavalin A which is also a member of the Diocleinae subtribe. However, differences in the magnitude of the thermodynamic binding parameters of the lectins are observed, even though the lectins possess conserved contact residues in many cases, and highly conserved primary sequences. The results indicate that non-contact residues in the lectins, even those distant from the binding sites, modulate their thermodynamic binding parameters.

Demonstration of a conserved histidine and two water ligands at the Mn2+ site in Diocleinae lectins by pulsed EPR spectroscopy.

Lectins from the Diocleinae subtribe, including Canavalia brasiliensis, Canavalia bonariensis, Canavalia grandiflora, Cratylia floribunda, Dioclea grandiflora, Dioclea guianensis, Dioclea rostrata, Dioclea violacea, and Dioclea virgata, have been recently isolated and characterized in terms of their carbohydrate binding specificities. Although all of the lectins are Man/Glc specific, they possess different biological activities. In the present study, electron paramagnetic resonance (EPR) spectroscopy demonstrates that all nine Diocleinae lectins contain Mn2+. The spectra of C. floribunda and D. rostrata suggest Mn2+ site symmetry different from that of the other seven lectins. However, electron spin-echo envelope modulation (ESEEM) spectroscopy indicates that all nine lectins are coordinated to a histidyl imidazole, with similar electron-nuclear coupling to the Mn2+-bound imidazole nitrogen. ESEEM also demonstrates ligation of two water molecules to Mn2+ in all nine Diocleinae lectins. Thus, the EPR and ESEEM data indicate the presence of a Mn2+ binding site in the above Diocleinae lectins with a conserved histidine residue and two water ligands.

Electron microscopy and x-ray diffraction studies of Lotus tetragonolobus A isolectin cross-linked with a divalent Lewisx oligosaccharide, an oncofetal antigen.

The interactions of lectins with multivalent carbohydrates often leads to the formation of highly ordered cross-linked lattices that are amenable to structural studies. A particularly well ordered, two-dimensional lattice is formed from fucose-specific isolectin A from Lotus tetragonolobus cross-linked with difucosyllacto-N-neohexaose, an oligosaccharide possessing the Lewisx determinant, which is an oncofetal antigen. A combination of electron microscopy, x-ray diffraction, simulation of electron micrographs, and molecular model building was used to determine the relative positions of the tetrameric lectin and bivalent carbohydrate within the lattice. X-ray diffraction from unoriented pellets was used to determine the lattice dimensions and analysis of electron micrographs was used to determine the lattice symmetry. Molecular models of the lattice were constructed based on the known structure of the jack bean lectin concanavalin A and the high degree of sequence homology between the two lectins. Using the symmetry and dimensions of the lattice and its appearance in filtered electron micrographs, molecular models were used to determine the orientation of the lectin in the lattice, and to define the range of lectin-oligosaccharide interactions consistent with the structural data. The present study provides the first description of a highly ordered, two-dimensional, cross-linked lattice between a tetravalent lectin and a bivalent carbohydrate.

Thermodynamics of binding of the core trimannoside of asparagine-linked carbohydrates and deoxy analogs to Dioclea grandiflora lectin.

The Man/Glc-specific seed lectin from Dioclea grandiflora (DGL) is a member of the Diocleinae subtribe that includes the jack bean lectin concanavalin A (ConA). Both DGL and ConA bind with high affinity to the "core" trimannoside moiety, 3, 6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranoside, which is present in asparagine-linked carbohydrates. Recent hemagglutination inhibition studies suggest that DGL and ConA recognize similar epitopes of the trisaccharide but possess different binding specificities for complex carbohydrates (Gupta, D., Oscarson, S., Raju, T. S., Stanley, P., Toone, E. J., and Brewer, C. F. (1996) Eur. J. Biochem. 242, 320-326). In the present study, we have used isothermal titration microcalorimetry to determine the thermodynamics of binding of DGL to a complete set of monodeoxy analogs of the core trimannoside as well as a tetradeoxy analog. The thermodynamic data indicate that DGL recognizes the 2-, 3-, 4-, and 6-hydroxyl groups of the alpha(1,6) Man residue, the 3- and 4-hydroxyl group of the alpha(1, 3) Man residue, and the 2- and 4-hydroxyl groups of the central Man residue of the trimannoside. The thermodynamic data for the tetradeoxy analog lacking the 3- and 4-hydroxyl group of the alpha(1, 3) Man residue, and the 2- and 4-hydroxyl groups of the central Man residue of the trimannoside are consistent with the involvement of these hydroxyl groups in binding. While the overall pattern of data for DGL binding to the deoxy analogs is similar to that for ConA (Gupta, D., Dam, T. K., Oscarson, S., and Brewer, C. F. (1997) J. Biol. Chem. 272, 6388-6392), differences exist in the data for certain monodeoxy analogs binding to the two lectins. Differences are also observed in the thermodynamics of binding of DGL and ConA to a biantennary complex carbohydrate. In the following paper (Rozwarski, D. A., Swami, B. M., Brewer, C. F., and Sacchettini, J. C. (1998) J. Biol. Chem. 273, 32818-32825), the x-ray crystal structure of DGL complexed to the core trimannoside is presented, and a comparison is made of the thermodynamic binding data for DGL and ConA as well as the structures of their respective trimannoside complexes.

Crystal structure of the lectin from Dioclea grandiflora complexed with core trimannoside of asparagine-linked carbohydrates.

The seed lectin from Dioclea grandiflora (DGL) has recently been shown to possess high affinity for 3, 6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranose, the core trimannoside of asparagine-linked carbohydrates, but lower affinity for biantennary complex carbohydrates. In the previous paper, the thermodynamics of DGL binding to deoxy analogs of the core trimannoside and to a biantennary complex carbohydrate were determined by isothermal titration microcalorimetry. The data suggest that DGL recognizes specific hydroxyl groups of the trimannoside similar to that of the jack bean lectin concanavalin A (ConA) (Gupta, D. Dam, T. K., Oscarson, S., and Brewer, C. F. (1997) J. Biol. Chem. 272, 6388-6392). However, the thermodynamics of DGL binding to certain deoxy analogs and to the complex carbohydrate are different from that of ConA. In the present paper, the x-ray crystal structure of DGL complexed to the core trimannoside was determined to a resolution of 2.6 A. The overall structure of the DGL complex is similar to the structure of the ConA-trimannoside complex (Naismith, J. H., and Field, R. A. (1996) J. Biol. Chem. 271, 972-976). The location and conformation of the bound trimannoside as well as its hydrogen-bonding interactions in both complexes are nearly identical. However, differences exist in the location of two loops outside of the respective binding sites containing residues 114-125 and 222-227. The latter residues affect the location of a network of hydrogen-bonded water molecules that interact with the trisaccharide. Differences in the arrangement of ordered water molecules in the binding site and/or protein conformational differences outside of the binding site may account for the differences in the thermodynamics of binding of the two lectins to deoxy analogs of the trimannoside. Molecular modeling studies suggest how DGL discriminates against binding the biantennary complex carbohydrate relative to ConA.

Differential solvation of "core" trimannoside complexes of the Dioclea grandiflora lectin and concanavalin A detected by primary solvent isotope effects in isothermal titration microcalorimetry.

The thermodynamics of binding of the Man/Glc-specific seed lectin from Dioclea grandiflora (DGL) to deoxy analogs of the "core" trimannoside, 3, 6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranoside was determined by isothermal titration microcalorimetry (ITC) in the first paper of this series (Dam, T. K., Oscarson, S., and Brewer, C. F. (1998) J. Biol. Chem. 273, 32812-32817). The data showed binding of specific hydroxyl groups on all three residues of the trimannoside, similar to that observed for ConA (Gupta, D., Dam, T. K., Oscarson, S., and Brewer, C. F. (1997) J. Biol. Chem. 272, 6388-6392). However, differences exist in the thermodynamics of binding of monodeoxy analogs of the alpha(1-6) Man residue of the trimannoside to the two lectins. The x-ray crystal structure of DGL complexed to the core trimannoside, presented in the second paper in this series (Rozwarski, D. A., Swami, B. M., Brewer, C. F., and Sacchettini, J. C. (1998) J. Biol. Chem. 273, 32818-32825), showed the overall structure of the complex to be similar to that of the ConA-trimannoside complex. Furthermore, the trimannoside is involved in nearly identical hydrogen bonding interactions in both complexes. However, differences were noted in the arrangement of ordered water molecules in the binding sites of the two lectins. The present study presents ITC measurements of DGL and ConA binding to the monodeoxy analogs of the trimannoside in hydrogen oxide (H2O) and deuterium oxide (D2O). The solvent isotope effects present in the thermodynamic binding data provide evidence for altered solvation of the parent trimannoside complexes at sites consistent with the x-ray crystal structures of both lectins. The results indicate that the differences in the thermodynamics of DGL and ConA binding to alpha(1-6) monodeoxy analogs of the trimannoside do not correlate with solvation differences of the parent trimannoside complexes.

Diocleinae lectins are a group of proteins with conserved binding sites for the core trimannoside of asparagine-linked oligosaccharides and differential specificities for complex carbohydrates.

The seed lectin from Dioclea grandiflora and jack bean lectin concanavalin A (ConA) are both members of the Diocleinae subtribe of Leguminosae lectins. Both lectins have recently been shown to possess enhanced affinities and extended binding sites for the trisaccharide, 3,6-di-O-(alpha-D-mannopyranosyl)-D-mannose, which is present in the core region of all asparagine-linked carbohydrates (Gupta, D., Oscarson, S., Raju, S., Stanley, P. Toone, E. J. and Brewer, C. F. (1996) Eur. J. Biochem. 242, 320-326). In the present study, the binding specificities of seven other lectins from the Diocleinae subtribe have been investigated by hemagglutination inhibition and isothermal titration microcalorimetry (ITC). The lectins are from Canavalia brasiliensis, Canavalia bonariensis, Cratylia floribunda, Dioclea rostrata, Dioclea virgata, Dioclea violacea, and Dioclea guianensis. Hemagglutination inhibition and ITC experiments show that all seven lectins are Man/Glc-specific and have high affinities for the core trimannoside, like ConA and D. grandiflora lectin. All seven lectins also exhibit the same pattern of binding to a series of monodeoxy analogs and a tetradeoxy analog of the trimannoside, similar to that of ConA and D. grandiflora lectin. However, C. bonariensis, C. floribunda, D. rostrata, and D. violacea, like D. grandiflora, show substantially reduced affinities for a biantennary complex carbohydrate with terminal GlcNAc residues, while C. brasiliensis, D. guianensis, and D. virgata, like ConA, exhibit affinities for the oligosaccharide comparable with that of the trimannoside. Thermodynamic data obtained by ITC indicate different energetic mechanisms of binding of the above two groups of lectins to the complex carbohydrate. The ability of the lectins to induce histamine release from rat peritoneal mast cells is shown to correlate with the relative affinities of the proteins for the biantennary carbohydrate.

Thermodynamics of lectin-carbohydrate interactions. Binding of the core trimannoside of asparagine-linked carbohydrates and deoxy analogs to concanavalin A.

The trisaccharide 3,6-di-O-(alpha-D-mannopyranosyl)-D-mannose, which is present in all asparagine-linked carbohydrates, was previously shown by titration microcalorimetry to bind to the lectin concanavalin A (ConA) with nearly -6 kcal mol-1 greater enthalpy change and 60-fold higher affinity than methyl-alpha-D-mannopyranoside (Mandal, D. K., Kishore, N., and Brewer, C. F. (1994) Biochemistry 33, 1149-1156). Similar studies of the binding of a series of monodeoxy derivatives of the alpha(1-3) residue of the trimannoside showed that this arm was required for high affinity binding (Mandal, D. K., Bhattacharyya, L., Koenig, S. H., Brown, R. D., III, Oscarson, S., and Brewer, C. F. (1994) Biochemistry 33, 1157-1162). In the present paper, a series of monodeoxy derivatives of the alpha(1-6) arm and "core" Man residue of the trimannoside as well as dideoxy and trideoxy analogs were synthesized. Isothermal titration microcalorimetry experiments establish that the 3-, 4-, and 6-hydroxyl groups of the alpha(1-6)Man residue of the trimannoside binds to the lectin, along with the 2- and 4-hydroxyl groups of the core Man residue and the 3- and 4-hydroxyl groups of the alpha(1-3)Man residue. Dideoxy analogs and trideoxy analogs showed losses of affinities and enthalpy values consistent with losses in binding of specific hydroxyl groups of the trimannoside. The free energy and enthalpy contributions to binding of individual hydroxyl groups of the trimannoside determined from the corresponding monodeoxy analogs are observed to be nonlinear, indicating differential contributions of the solvent and protein to the thermodynamics of binding of the analogs. The thermodynamic solution data agree well with the recent x-ray crystal structure of ConA complexed with the trimannoside (Naismith, J. H., and Field, R. A. (1996) J. Biol. Chem. 271, 972-976).

X-ray crystallographic studies of unique cross-linked lattices between four isomeric biantennary oligosaccharides and soybean agglutinin.

Soybean agglutinin (SBA) (Glycine max) is a tetrameric GalNAc/Gal-specific lectin which forms unique cross-linked complexes with a series of naturally occurring and synthetic multiantennary carbohydrates with terminal GalNAc or Gal residues [Gupta et al. (1994) Biochemistry 33, 7495-7504]. We recently reported the X-ray crystal structure of SBA cross-linked with a biantennary analog of the blood group I carbohydrate antigen [Dessen et al. (1995) Biochemistry 34, 4933-4942]. In order to determine the molecular basis of different carbohydrate-lectin cross-linked lattices, a comparison has been made of the X-ray crystallographic structures of SBA cross-linked with four isomeric analogs of the biantennary blood group I carbohydrate antigen. The four pentasaccharides possess the common structure of (beta-LacNAc)2Gal-beta-R, where R is -O(CH2)5COOCH3. The beta-LacNAc moieties in the four carbohydrates are linked to the 2,3-, 2,4-, 3,6-, and 2,6-positions of the core Gal residue(s), respectively. The structures of all four complexes have been refined to approximately 2.4-2.8 A. Noncovalent lattice formation in all four complexes is promoted uniquely by the bridging action of the two arms of each bivalent carbohydrate. Association between SBA tetramers involves binding of the terminal Gal residues of the pentasaccharides at identical sites in each monomer, with the sugar(s) cross-linking to a symmetry-related neighbor molecule. While the 2,4-, 3,6-, and 2,6-pentasaccharide complexes possess a common P6422 space group, their unit cell dimensions differ. The 2, 3-pentasaccharide cross-linked complex, on the other hand, possesses the space group I4122. Thus, all four complexes are crystallographically distinct. The four cross-linking carbohydrates are in similar conformations, possessing a pseudo-2-fold axis of symmetry which lies on a crystallographic 2-fold axis of symmetry in each lattice. In the case of the 3,6- and 2,6-pentasaccharides, the symmetry of their cross-linked lattices requires different rotamer orientations about their beta(1,6) glycosidic bonds. The results demonstrate that crystal packing interactions are the molecular basis for the formation of distinct cross-linked lattices between SBA and four isomeric pentasaccharides. The present findings are discussed in terms of lectins forming unique cross-linked complexes with glycoconjugate receptors in biological systems.

Thermodynamics of carbohydrate binding to galectin-1 from Chinese hamster ovary cells and two mutants. A comparison with four galactose-specific plant lectins.

The thermodynamics of carbohydrate binding to the 14 kDa dimeric beta-galactoside-binding lectin galectin-1 (Gal-1) from Chinese hamster ovary cells and four galactose-specific plant lectins were investigated by isothermal titration microcalorimetry. Recombinant Gal-1 from Escherichia coli, a Cys-->Ser mutant with enhanced stability (C2S-Gal-1), and a monomeric mutant of the lectin (N-Gal-1) were studied along with the soybean agglutinin and the lectins from Erythrina indica, Erythrina crystagalli, and Erythrina corollodendrum. Although the pattern of association constants of the Erythrina lectins was similar for mono- and disaccharides, variations exist in their enthalpy of binding (-delta H) values for individual carbohydrates. While the Erythrina lectins show greater affinities and -delta H values for lactose and N-acetyllactosamine, the soybean agglutinin possesses similar affinities for methyl beta-galactopyranoside, lactose, and N-acetyllactosamine and a greater -delta H value for the monosaccharide. Gal-1 and the plant lectins possess essentially the same affinities for N-acetyllactosamine; however, the animal lectin shows a lower -delta H value and more favorable binding entropy for the disaccharide. While Gal-1, C2S-Gal-1, and N-Gal-1 all possess essentially the same affinities for N-acetyllactosamine, the two mutants possess much lower -delta H values, even though the mutation site(s) are far removed from the carbohydrate binding site. These results indicate that there are different energetic mechanisms of carbohydrate binding between galectin-1, its two mutants, and the Gal-specific plant lectins.

A comparison of the fine saccharide-binding specificity of Dioclea grandiflora lectin and concanavalin A.

The lectin from the seeds of Dioclea grandiflora (DGL) is a Man/Glc-specific tetrameric protein with physical and saccharide-binding properties reported to be similar to that of the jack bean lectin concanavalin A (ConA). Unlike other plant lectins, both DGL and ConA bind with high affinity to the core trimannoside moiety, 3,6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranoside, which is present in all asparagine-linked carbohydrates. In the present study, hemagglutination inhibition techniques have been used to investigate binding of DGL and ConA to a series of mono- and dideoxy analogs of methyl 3,6-di-O-(alpha-D-mannopyranosyl)-alpha-D-mannopyranoside and to a series of asparagine-linked oligomannose and complex oligosaccharides and glycopeptides. The results indicate that both DGL and ConA recognize epitopes on all three residues of the trimannoside: the 3-, 4-, and 6-hydroxyl groups of the alpha(1-6)Man residue, the 3-hydroxyl group of the alpha(1-3)Man residue, and the 2- and 4-hydroxyl groups of the central Man residue of the core trimannoside. However, unlike ConA, DGL does not bind to biantennary complex carbohydrates. This was confirmed by showing that biantennary complex glycopeptides do not bind to a DGL-Sepharose affinity column. Unlike ConA, DGL does not show enhanced affinity for a large N-linked oligomannose carbohydrate (Man9 glycopeptide) relative to the trimannoside. Thus, DGL and ConA share similar epitope recognition of the core trimannoside moiety. However, they exhibit differences in their fine specificities for larger N-linked oligomannose and complex carbohydrates.