Stereochemical metrics of lectin-carbohydrate interactions: comparison with protein-protein interfaces

A global census of stereochemical metrics including interface size, hydropathy, amino acid propensities, packing and hydrogen bonding was carried out on 32 x-ray-elucidated structures of lectin-carbohydrate complexes covering eight different lectin families. It is shown that the interactions at primary binding subsites are more efficient than at other subsites. Another salient behavior found for primary subsites was a marked negative correlation between the interface size and the polar surface content. It is noteworthy that this demographic rule is delineated by lectins with unrelated phylogenetic origin, indicating that independent interface architectures have evolved through common optimization paths. The structural properties of lectin-carbohydrate interfaces were compared with those characterizing a set of 32 protein homodimers. Overall, the analysis shows that the stereochemical bases of lectin-carbohydrate and protein-protein interfaces differ drastically from each other. In comparison with protein-protein complexes, lectin-carbohydrate interfaces have superior packing efficiency, better hydrogen bonding stereochemistry, and higher interaction cooperativity. A similar conclusion holds in the comparison with protein-protein heterocomplexes. We propose that the energetic consequence of this better interaction geometry is a larger decrease in free energy per unit of area buried, feature that enables lectins and carbohydrates to form stable complexes with relatively small interface areas. These observations lend support to the emerging notion that systems differing from each other in their stereochemical metrics may rely on different energetic bases.     

Effect of a specific inhibitor on the unfolding and refolding kinetics of dimeric triosephosphate isomerase: establishing the dimeric and similarly structured nature of the main transition states on the forward and backward reactions

2-Phosphoglycolate (PGA), a strong competitive inhibitor of the dimeric enzyme triosephosphate isomerase (TIM), brings about a large decrease in the unfolding rate constant of the protein. The data set of rate constants versus ligand concentration may be equally well explained by regarding either a monomeric or a dimeric transition state (TS). However, if the thermodynamics for binding of PGA to native TIM is taken into account, it becomes clear that a dimeric TS is the right assumption. Furthermore, by studying the effect of the ligand on the second-order refolding reaction, we found results indicating similar PGA-binding affinities to be present in the transition states for the rate-limiting steps of the forward and backward reactions. Most likely, therefore, both TS resemble each other in respect to the active site architecture. It should be mentioned, however, that our data do not rule out the possible occurrence of an unstable, (partially) folded monomeric intermediate, which would rapidly interconvert with the unfolded monomer.     

Bioavailability Studies of Stabilized Iron (II) Sulfate by Means of the Prophylactic-preventive Method

The bioavailability of stabilized ferrous sulfate was studied by means of the prophylactic-preventive test in rats. For comparative purposes, ferrous sulfate was used as reference standard. The test was performed in male weaned rats during 3 weeks, which were randomized into three groups of ten animals each. A control group received a basal diet of low iron content while the other groups received the same diet added with iron at a dose of 15 mg/kg as FeSO₄ 7H₂O and stabilized ferrous sulfate, respectively. Individual hemoglobin concentrations and weights were determined at the beginning and at the end of the study, and food intake was daily registered. Iron bioavailability (BioFe) of each source was calculated as the ratio between the amount of iron incorporated into hemoglobin during the treatment and the total iron intake per animal. A relative biological value was obtained as the ratio between the BioFe of stabilized ferrous sulfate and the reference standard given a value of 96%. Stabilized ferrous sulfate showed a high bioavailability, and when it is used to fortify dairy products as cheese and fluid milk in a dose of 15–20 mg of iron per kilogram, it behaved inertly in relation to the sensorial properties of the fortified food. These results suggest that this iron compound is a promising source to be use in food fortification.     

Posters

Introduction  Fine needle aspiration (FNA) cytology of the thyroid is a well-established test in the clinical work-up of patients with solitary nodules of the thyroid. Thyroid FNA does however have limitations and audit of diagnostic performance is important.
Methods  The histopathology archives of the Royal Victoria Hospital were searched for all thyroid resections and the histopathological diagnosis was correlated with the pre-operative cytological diagnosis, where available. Special emphasis was placed on the accuracy of tumour diagnosis.
Results  A total of 173 cases were identified during the 2-year period, of these 93 had available pre-operative FNA. A total of 57 tumours were identified. A small number (six of 57) of significant discrepancies were identified. These included a malignant lymphoma diagnosed as Hashimoto's thyroiditis, a metastasis which the FNA had suggested was a medullary carcinoma and an insular carcinoma diagnosed as medullary carcinoma on FNA. False positives included a colloid cyst diagnosed as suspicious of malignancy and a cytological diagnosis of papillary carcinoma not confirmed on histology.
Discussion  At present, the majority of thyroid FNAs in our clinics are performed by surgeons and material is not routinely available for immunocytochemistry. In spite of these limitations, there were few major discrepancies. These might be reduced if pathologist aspirators were able to perform FNAs and collect material for further studies, where necessary. This would allow identification of medullary carcinomas and malignant lymphomas.
Conclusion  FNA of thyroid lesions is a useful investigation in our clinical setting, however, some areas of potential for improvement have been identified.     

Stabilized ferrous gluconate as iron source for food fortification

The iron bioavailability and acute oral toxicity in rats of a ferrous gluconate compound stabilized with glycine (SFG), designed for food fortification, was studied in this work by means of the prophylactic method and the Wilcoxon method, respectively. For the former studies, SFG was homogenously added to a basal diet of low iron content, reaching a final iron concentration of 20.1±2.4 mg Fe/kg diet. A reference standard diet using ferrous sulfate as an iron-fortifying source (19.0±2.1 mg Fe/kg diet) and a control diet without iron additions (9.3±1.4 mg Fe/kg diet) were prepared in the laboratory in a similar way. These diets were administered to three different groups of weaning rats during 23 d as the only type of solid nourishment. The iron bioavailability of SFG was calculated as the relationship between the mass of iron incorporated into hemoglobin during the treatment and the total iron intake per animal. This parameter resulted in 36.6±6.2% SFG, whereas a value of 35.4±8.0% was obtained for ferrous sulfate. The acute toxicological studies were performed in two groups of 70 female and 70 male Sprague-Dawley rats that were administered increasing doses of iron from SFG. The LD₅₀ values of 1775 and 1831 mg SFG/kg body wt were obtained for female and male rats, respectively, evidencing that SFG can be considered as a safe compound from a toxicological point of view.     

Proffered papers 9.30–10.00 Monday 15 September 2003

Both the original Bethesda system and the current UK classifications of cervical cytology have proved robust but each has a major weakness in the area of abnormalities of uncertain significance. Cytologists recognize that sometimes it is simply impossible to differentiate between reactive and dyskaryotic material. For this reason, the Australian version of the Bethesda system introduced a new category of 'high grade inconclusive' with a recommendation for referral to colposcopy. Approximately 60% of such cases are found to have high grade lesions at colposcopy (Schoolland M, Sterrett G, Knowles S et al .). The present UK system even with the proposed changes requires of the pathologist, a decision as to whether such cases are probably high grade (=a report of moderate dyskaryosis) or not (= a report of borderline). This continues to ignore the fact that sometimes you just cannot tell, even on review. We have taken a consecutive series of 50 referral smears, reported as moderate dyskaryosis, where the histological outcome (by loop cone) is known. These cases were rescreened and then reviewed blind by a pathologist with extensive experience of the Australian NH & MRC modified Bethesda system. On review, the material was reclassified along NH & MRC lines. The results were compared with the biopsy findings in order to determine whether the category of 'inconclusive' might be of value in the context of the NHSCSP.     

POTENTIAL OF DISCARIA AMERICANA FOR METAL STABILIZATION ON SOILS AMENDED WITH BIOSOLIDS AND ASH-SPIKED BIOSOLIDS

Biosolids (B) may contain various types of environmental pollutants, which can exert phytotoxic effects in plants. The effect of aqueous extracts on seed germination and the primary root growth of discaria (Discaria americana) obtained from different soil-application rates of B and a mixture of B and incinerated B were investigated. The objective was to evaluate the potential use of discaria for the stabilization of B-amended soils. Ryegrass (Lolium perenne L.) was used for comparison. Compared to ryegrass, relative seed germination (RSG) was significantly lower for discaria. RSG of discaria and rye grass was inversely correlated to the electrical conductivity of extracts, although a significant adverse effect was only observed for ryegrass with the highest dose of the mixture of B and incinerated B. This dose also produced a reduction in the germination index of discaria, which could not be correlated with the parameters studied. The B extracts did not exert any significant adverse effect on the relative root growth of both species. An increase in relative root growth and germination index was observed for discaria with a field application rate equivalent of 156 t DW ha^?1 of B, suggesting a stimulating effect of the amendment. The results obtained in this study suggest that germinated seedlings of discaria might be used for the stabilization of B-amended soils. However, further greenhouse and field experiments should be performed to confirm these findings.