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Intrafamily gene diversification has led to three prototype galectins in chicken [i.e., chicken galectin (CG)-1A, CG-1B, and CG-2] that show distinct expression profiles and developmental regulation. In order to pinpoint structural disparities among them, we determined the crystal structure of CG-1B. Alteration of the position of the Trp ring in the lectin site and the presence of only two ordered water molecules therein, as well as changes in the interface region between the two subunits, set the structure of CG-1B clearly apart from that of CG-1A. Intriguingly, the unique presence of two Cys residues at positions 2 and 7 in the N-terminal region translated into formation of an intersubunit disulfide bridge between the Cys7 residues of the homodimer in the crystal. In solution, oxidation is associated with significant shape changes in the dimeric protein and the additional occurrence of a compacted form with an intrasubunit disulfide bridge between Cys2 and Cys7. The single-site mutant C7S/C7V was not subjected to such changes, supporting the crucial role of Cys7 in redox-dependent shape changes. These results point to the functional significance of the distinctive presence of the two Cys residues in the N-terminal region of CG-1B.

Glycans of natural glycoconjugates are considered as a source of biological information relevant to cell adhesion or growth. Sugar-based messages are decoded and translated into responses by endogenous lectins. This mechanism assigns a functional dimension to tumour-associated changes of glycosylation. Consequently, it calls for mapping the lectin presence in tumours. Such an analysis has so far commonly been performed with the scope to determine expression of a few distinct proteins, e.g. from the effector family of galectins with focus on galectins-1 and -3. Due to the emerging evidence for functional divergence among galectins it is timely to address the challenge to evaluate their presence beyond these few family members. Having raised a panel of non-cross- -reactive antibodies against seven human galectins covering all three subfamilies, we de scribe their expression profiles in human skin. Comparison of normal and malignant tissues enabled us to define galectin-type-dependent alterations, arguing in favour of distinct functionalities. It is concluded that comprehensive monitoring performed to define the different aspects of the galectin network, as documented in this pilot study, is advisable for future histopathologic studies aimed at delineating clinical correlations.

Gene duplication and sequence divergence are driving forces toward establishing protein families. To examine how sequence changes affect carbohydrate specificity, the two closely related proto-type chicken galectins CG-14 and CG-16 were selected as models. Binding properties were analyzed using a highly sensitive solid-phase assay. We tested 56 free saccharides and 34 well-defined glycoproteins. The two galectins share preference for the II (Galbeta1-4GlcNAc) versus I (Galbeta1-3GlcNAc) version of beta-galactosides. A pronounced difference is found owing to the reactivity of CG-14 with histo-blood group ABH active oligosaccharides and A/B active glycoproteins. These experimental results prompted to determine activity-structure correlations by modeling. Computational analysis included consideration of the flexibility of binding partners and the presence of water molecules. It provided a comparative description of complete carbohydrate recognition domains, which had so far not been characterized in animal galectins. The structural models assigned II, I selectivity to a region downstream of the central Trp moiety. Docking revealed that the tetrasaccharides can be accommodated in their free-state low-energy conformations. CG-14's preference for A versus B epitopes could be attributed to a contact between His124 and the N-acetyl group of GalNAc. Regarding intergalectin comparison, the Ala53/Cys51 exchange affects the interaction potential of His54/His52. Close inspection of simulated dynamic interplay revealed reorientation of His124 at the site of the His124/Glu123 substitution, with potential impact on ligand dissociation. In summary, this study identifies activity differences and provides information on their relation to structural divergence, epitomizing the value of this combined approach beyond galectins.

Presence of species-specific gene divergence in a protein family prompts to thoroughly study structural aspects and expression profiles of the products. We herein focus on two members of an adhesion/growth-regulatory group of endogenous lectins, i.e. galectins-5 and -9. After first ascertaining species specificity of occurrence of galectin-5, constituted by a short section of rat galectin-9's N-terminal part and its C-terminal carbohydrate recognition domain, by database mining, we next detected and defined sequence differences in the proximal promoter region between the two genes. The ensuing hypothesis for distinct expression profiles was tested first by RT-PCR and then by immunohistochemistry. For the latter purpose, we employed antibodies rigorously controlled for absence of cross-reactivity including assays with various other galectins and, if necessary, refined by chromatographic removal of bi- or oligospecific activities. Indeed, the galectins have non-identical expression profiles, qualitative differences, e.g. seen for galectin-5-positive bone marrow and erythrocytes or for hitherto unknown expression in cells of the theca folliculi and galectin-9-positive skin epidermis and esophageal epithelium. Lack of hepatocyte or renal cortex staining separates these two expression profiles in rat from localization of galectin-9 in mouse. Interspecies extrapolation in a case of a galectin involved in unique gene divergence may thus not be valid. The presented results on galectin-5 relative to galectin-9 intimate distinct functions especially in erythropoiesis and imply currently unknown mechanisms to compensate its absence from the galectin network in other mammals.

The growth-regulatory interplay between ganglioside GM1 on human SK-N-MC neuroblastoma cells and an endogenous lectin provides a telling example for glycan (polysaccharide) functionality. Galectin-1 is the essential link between the sugar signal and the intracellular response. The emerging intrafamily complexity of galectins raises the question on defining extent of their structural and functional overlap/divergence. We address this problem for proto-type galectins in this system: ganglioside GM1 as ligand, neuroblastoma cells as target. Using the way human galectin-1 interacts with this complex natural ligand as template, we first defined equivalent positioning for distinct substitutions in the other tested proto-type galectins, e.g., Lys63 vs. Leu60/Gln72 in galectins-2 and -5. As predicted from our in silico work, the tested proto-type galectins have affinity for the pentasaccharide of ganglioside GM1. In contrast to solid-phase assays, cell surface presentation of the ganglioside did not support binding of galectin-5, revealing the first level of regulation. Next, a monomeric proto-type galectin (CG-14) can impair galectin-1-dependent negative growth control by competitively blocking access to the shared ligand without acting as effector. Thus, the quaternary structure of proto-type galectins is an efficient means to give rise to functional divergence. The identification of this second level of regulation is relevant for diagnostic monitoring. It might be exploited therapeutically by producing galectin variants tailored to interfere with galectin activities associated with the malignant phenotype. Moreover, the given strategy for comparative computational analysis of extended binding sites has implications for the rational design of galectin-type-specific ligands.

Members of the galectin family of endogenous lectins are involved in tumor growth regulation and in establishing characteristics of the malignant phenotype via protein-carbohydrate and protein-protein interactions. To identify peptide ligands with the potential to modulate these tumor-relevant interactions beneficially, complementary screening methods were employed, that is, both phage-display and a combinatorial pentapeptide library with the key YXY tripeptide core. Three representative prototype galectins were selected. The search for high-affinity ligands among phage-displayed random heptamers yielded enrichment after five selection cycles of the nonglycomimetic CQSPSARSC peptide in the case of the chicken homologue of galectin-1 but not the human protein, an indication for specificity. The most active glycomimetic from the combinatorial library of 5832 pentamers was WYKYW. Identification of peptide ligands for galectins with and without glycomimetic properties is thus possible. Our study documents the potential to combine the two library-based approaches for structural optimization of lead peptides.

Membrane glycoconjugates play a central role in neuronal interactions and regulation. To define the precise links between membrane polysaccharides and neuronal functions, two main requirements must be fulfilled: (1) the availability of molecular tools able to finely discriminate among carbohydrate structures and (2) the use of an experimental system suitable for systematic and quantitative studies of particular neuronal processes. In this work, we used two chicken proto-type galectins, i.e., monomeric CG-14 and dimeric CG-16, with very similar carbohydrate affinities, and rat hippocampal neurons in culture to quantitatively measure the involvement of carbohydrate-protein interaction in axonal growth and directionality, neurite sprouting and axon regenerative capacity after section. CG-16 potently stimulated axonal growth and guidance. Neurite sprouting was enhanced by immobilized CG-16 and, notably, reduced by lectin in solution. Overall, cross-linking CG-16 invariably excelled CG-14 in these functional assays, although none of them were able to improve axon regenerative capacity when compared to mammalian galectin-1. Our results demonstrate the potential of the experimental set-up to perform a systematic study of galectin functionality in neuronal differentiation. In view of the concept of the sugar code, the presented results indicate that biological effects triggered by glycan binding engaging an endogenous lectin can be modulated by carbohydrate affinity and/or by other factors like differential cross-linking capacity.

A library of pentapeptides containing the sequence -Y-X-Y- based on rational design was screened with six different lectins. Sequences were identified that modulate galectin binding to its natural carbohydrate ligand. SPR showed inhibition values 2-3 times stronger than galactose and NMR studies suggested real carbohydrate mimicry.

The genes nikQ and nikR were identified by sequencing DNA of the nikkomycin biosynthetic gene cluster from Streptomyces tendae Tü901/8c. The nikQ gene encodes a P450 cytochrome, and the predicted NikR gene product shows 48-56% sequence identity with uracil phosphoribosyltransferases from eukaryotic organisms. The nikQ and nikR genes were inactivated separately by insertion of a kanamycin-resistance cassette. Inactivation of the nikQ gene abolished synthesis of nikkomycins containing 4-formyl-4-imidazolin-2-one as the base (nikkomycins X and I), whereas production of nikkomycins containing uracil (nikkomycins Z and J) was not affected. Nikkomycin X and I production could be restored by feeding 4-formyl-4-imidazolin-2-one to the nikQ mutants, indicating that NikQ is responsible for its formation from L-histidine. Disruption of the nikR gene resulted in formation of decreased amounts of nikkomycins X and I, whereas nikkomycins Z and J were synthesized at wild-type levels. A fluorouracil-resistant nikR mutant lacking uracil phosphoribosyltransferase (UPRTase) activity did not synthesize nikkomycins X and I and accumulated 4-formyl-4-imidazolin-2-one in its culture filtrate, whereas formation of nikkomycins Z and J was unimpaired. The mutant was complemented to nikkomycin X and I production by nikR expressed from the mel promoter of plasmid pIJ702. The nikR gene expressed in Escherichia coli led to the production of UPRTase activity. Our results indicate that NikR converts 4-formyl-4-imidazolin-2-one to yield 5'-phosphoribosyl-4-formyl-4-imidazolin-2-one, the precursor of nikkomycins containing this base.

The Torre-Muir syndrome is a tumour syndrome characterized by multiple cutaneous and extracutaneous primary carcinomas and different sebaceous neoplasias. The visceral adenocarcinomas are mainly found in the gastrointestinal and urogenital tract. A case of Torre-Muir syndrome is described in a female patient and the different tumour localizations are demonstrated.

The effects of Candida albicans on the healing of experimental wounds in rats was investigated. Granulation tissue developing in implated polyvinylchloride chambers was quantified. The amount of granulation tissue in the yeast-contaminated wounds is significantly less in comparison to controls. The inhibitory effect is directly proprotional to the amount of inoculated yeasts and dependes on the presence of living yeast cells. The repair of ulcers is supposedly retarded by yeast contamination, as has been suggested previously by clinical observation.

Erythema chronicum migrans (E.c.m.), Acrodermatitis chronica atrophicans (A.c.a.) and Lymphadenosis benigna cutis (L.b.c.) are infectious diseases according to Koch's criteria, after the infectious agent-Borrelia-Burgdorferi-could could be isolated. Clinic, pathology and therapy of these three diseases are presented in detail. The correlation to the lyme disease is being discussed. We believe that E.c.m., A.c.a. and L.b.c. are different entities determined by the virulence of the infectious agent and the individual immunological response.

The case report of a 83-year-old woman with lichen ruber ulcerosus is reviewed. Clinically the patient showed extensive ulcerations of the sole of one foot, with synechiae of the toes. In addition, there was subtotal atrophic alopecia of the scalp, vulvar atrophy, and circumscribed cutaneous lesions of lichen planus. The disease started with chronic recurrent paronychiae. The ulcerative defect was successfully treated with mesh grafting, which is the therapy of choice.

A lot of substances currently used in medicine and daily life are found to induce pseudoallergic reactions. Because of practical importance intolerance to aspirin and additives as well as anaphylactoid reactions to sulfites and local anaesthetics are presented. The aetiopathological problems are not yet solved, and the differentiation from true allergic diseases is difficult. Concerning the local anaesthetics, additional side effects of other components of the solutions have to be considered. An accurate history is often worthy. Generally, it is not possible to clear up pseudoallergic reactions through skin tests, so that incremental challenge is necessary.