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The N epsilon-glycosylation of lysine and hydroxylysine residues in collagen from streptozotocin-induced-diabetic rats was confirmed and the stability of the complex shown to be due to an Amadori rearrangement. The studies also demonstrate the relative specificities of glucose, galactose and mannose in their reaction with collagen. The glycosylation of lysine in vitro occurs with glucose and galactose, but not with mannose, whereas only gucose reacts with hydroxylysine to any significant extent. Glycosylation of collagen occurs slowly during normal aging, but in contrast with reports suggesting accelerated aging of collagen in diabetic animals, we clearly demonstrated that the apparent increased stability is not due to an acceleration of the normal maturation process involving the reducible cross-links.

Acid-soluble collagen from rat tail tendon was glucosylated in vitro and fibrillogenesis parameters were determined first at 35 degrees C then at 4 degrees C. Increased lag phase and half time were shown to be related to the amount of non-enzymatically bound glucose, probably due to a decrease of hydrophobic interactions at this early stage of fibril formation. The absence of intermolecular cross-links and the partial redissolution of fibrils of 4 degrees C, as investigated both by turbidimetry and electron microscopy, suggests a defect in the maturation process in glucosylated collagen fibrils.

Summary This study proposes to show the possible relationships between, on the one hand, modifications of evoked potentials (E.P.) and slow potentials (S.P.) and, on the other hand anomalies in amino acid metabolism in mentally deficient children. Thirty-two mentally retarded children (I. Q.'s between 20 and 40) were examined and divided into two groups : R1 with no amino acid anomalies ; R2 with amino acid anomalies, « isolatedor associated with phosphocalcium metabolism perturbations or renal dysfunction. As predicted by previous observations, these mentally deficient children presented low amplitude E.P. and S.P. irrespective of their group assignment. Both pathological groups are characterised by a lack of E. P. conditioning. Moderate differences are observed between the two groups, R1 and R2. These differences are most evident in E.P. amplitude and less evident in S.P. These responses are weaker in group R2 than in R1. Within group R2 they are also weaker for those children with isolated amino acide anomalies. The low amplitude of evoked potentials observed in mentally retarded children can be linked, not only to perceptive and cognitive factors, but to biological factors.

Ubiquitin was extracted from testis of 4 mammals and purified to homogeneity by gel filtration chromatography. Amino acid compositions and NH2-terminal sequences were found to be identical in the 4 species and with calf thymus ubiquitin. Ubiquitin conformation was shown to be very sensitive to oxidation. Improved methods for radioimmunoassay of ubiquitin in tissue extracts are also discussed.

A technique of preparative isotachophoresis is described for the preparation of two rat α-1-macroglobulin varieties. The procedure permits a good and fast separation. Antigenic identity of these components is shown by fused rocket immunoelectrophoresis. Some properties of the two varieties are determined, and causes of heterogeneity are also discussed.

Four crude glycoproteins were isolated from supernatant of a Wistar rat kidney cortex homogenate. Three were soluble in a saline solution at pH 7.5; a fourth was prepared from a trypsin hydrolysate. T

Glomerular basement membranes were isolated from normal and streptozotocin-diabetic rats. after elimination of non collagenous components with 8M urea the extent of ketoamine-linked hexoses and cross-links in the preparation was determined using NaB 3 H 4 reduction and radiochromatography. The observed increase in the level of N-glycosylation appeared to be closely related to a lowered amount of intermolecular cross-links in diabetic rats as compared to normal animals. Such a glycosylation may affect the stability and physicochemical properties of G.B.M. and thereby contribute to the vascular sequelae of diabetes mellitus.

1. 1. The study of the amino acid composition, in particular the study of the hydroxyproline, hydroxylysine and glycine content, of the glycopeptide fractions obtained from a clostridropeptidase A hydrolysate of Wistar rat GBM allowed us to distinguish between two types of structure, one of which is collagen-like. 2. 2. Determination of carbohydrate composition revealed the existence of a heteropolysaccharidic glycane in all of the fractions studied. 3. 3. The study of the chemical composition of the high molecular weight components isolated using a Sepharose 4B column allowed us to distinguish between structural glycoproteins (Subfractions A 1 , A 2 , A 4 ) and collagen-like glycopeptides (Subfraction A 3 ) to which they are linked. 4. 4. The immunological studies revealed the antigenic complexity of the material studied and the existence of precipitin lines of identity for the fractions which were isolated.

Non-enzymatic glycosylation of collagen occurs both in vivo during diabetes and in vitro after incubation with glucose. Glycosylated collagen exhibits altered physicochemical and biological properties which could explain some of the complications of diabetes. To provide a mechanistic explanation of this modification the localization of bound glucose was investigated using NaB[3H]H4 reduction and CNBr cleavage. Glucose fixation is distributed mainly on the alpha 1CB6 peptide after in vitro glycosylation whereas this distribution occurs less specifically during diabetes. It is concluded that fibrillogenesis alteration of in vitro glycosylated collagen is related to glucose fixation on free epsilon NH2 sites normally implied in intermolecular interactions.

Kinetic studies of complement activation were followed by hemolytic assay. Mathematical analysis shows that the curve is composed of two exponents: the first one, which occurs during a short span of time, represents the classical pathway, the second the alternative pathway. We were therefore able to foretell the respective participation of each activator used: inulin, zymosan, and aggregated immunoglobulins.

The effect of normal and nonenzymatically glycosylated rat type I acid-soluble collagen on normal human platelets was investigated. Glycosylated collagen was obtained either after in vitro incubation with glucose or from rats made diabetic by streptozotocin. The amount of nonenzymatically bound glucose was as follows: normal 2.3, diabetic 7.6 and in vitro glycosylated collagen 9.0 nmol/mg. When investigated under conditions leading to identical diameters and molecular packings for all the samples, the aggregation potency was markedly stronger for diabetic and glycosylated collagens than for normal ones. These results show the potential role of this posttranslational modification of collagen which can be considered as a risk factor in the thrombotic pathogeny of diabetes.

In vitro non-enzymatic condensation of glucose on acid soluble collagen is shown not to affect the molecular structure and stability as evaluated both by circular dichroism and differential spectrometry. Viscometric studies demonstrate a lowering in intermolecular interactions; whereas the hydrodynamic volume remains unaffected by nonenzymatic glycosylation. alpha 2(I) collagen chains exhibit a modified electrophoretic mobility previously found in collagen of diabetic rats; this peculiar behavior is discussed in terms of lowered hydrophobicity due to addition of hydrophilic groups from glucose. The lowered formation of hydrophobic areas, evaluated from the fluorescence emission of diphenylhexatriene, indicates that non-enzymatic glycosylation is able to influence the close packing of collagen fibers.