Characterization of Glycated Proteins by \u3csup\u3e13\u3c/sup\u3eC NMR Spectroscopy

13C NMR spectroscopy has been used to characterize Amadori (ketoamine) adducts formed by reaction of [2-13C]glucose with free amino groups of protein. The spectra of glycated proteins were acquired in phosphate buffer at pH 7.4 and were interpreted by reference to the spectra of model compounds, N alpha-formyl-N epsilon-fructose-lysine and glycated poly-L-lysine (GlcPLL). The anomeric carbon region of the spectrum (approximately 90-105 ppm) of glycated cytochrome c was superimposable on that of N alpha-formyl-N epsilon-fructose-lysine, and contained three peaks characteristic of the alpha- and beta-furanose and beta-pyranose anomers of Amadori adducts to peripheral lysine residues on protein (pK alpha approximately 10.5). The spectrum of GlcPLL yielded six anomeric carbon resonances; the second set of three was displaced about 2 ppm to lower shielding of the first and was assigned to the Amadori adduct at the alpha-amino terminus (pK alpha approximately 7.5). The spectrum of glycated RNase was similar to that of GlcPLL, but contained a third set of three signals attributable to modification of active site lysine 41 (pK alpha approximately 8.8). The assignments for RNase were confirmed by analysis of spectra taken at pH 4 and under denaturing conditions. The spectrum of glycated hemoglobin was comparable to that of GlcPLL, and distinct resonances could be assigned to Amadori adducts at amino-terminal valine and intrachain N epsilon-lysine residues. Chemical analyses were performed to measure the relative extent of alpha- and epsilon-amino group modification in the glycated macromolecules, and the results were compared with estimates based on integration of the NMR spectra

GRADES: Gradient descent for similarity caching

A similarity cache can reply to a query for an object with similar objects stored locally. In some applications of similarity caches, queries and objects are naturally represented as points in a continuous space. Examples include 360° videos where user's head orientation - expressed in spherical coordinates - determines what part of the video needs to be retrieved, and recommendation systems where the objects are embedded in a finite-dimensional space with a distance metric to capture content dissimilarity. Existing similarity caching policies are simple modifications of classic policies like LRU, LFU, and qLRU and ignore the continuous nature of the space where objects are embedded. In this paper, we propose Grades, a new similarity caching policy that uses gradient descent to navigate the continuous space and find the optimal objects to store in the cache. We provide theoretical convergence guarantees and show Grades increases the similarity of the objects served by the cache in both applications mentioned above

Influence of days after calving and thermal stress on the efficacy of a progesterone-based treatment in acyclic italian mediterranean buffalo

The aim of this study was to evaluate the efficacy of a progesterone-based treatment on anoestrus in buffaloes. Primiparous acyclic buffaloes (n = 276), were divided into three classes according to their days in milk (DIM): from 50 to 90 (Class I; n = 86), from 91 to 150 (Class II; n = 102) and from 150 to 200 (Class III; n = 88). Animals were synchronized using P4 vaginal implants, followed by timed artificial insemination (TAI). They were then allowed to enter into a larger group of buffaloes for natural mating 15 days after AI was performed, and pregnancy status was monitored from then on at 15-day intervals. Finally, the temperature–humidity index (THI) was calculated. Statistical analysis was performed by ANOVA by means and both multiple and linear regression. The total pregnancy rate (PR) was 87.7%, with no differences among DIM classes (88.0, 92.4, and 80.0% in Classes I, II, and III, respectively). However, the PR at TAI tended to be higher (p = 0.07) in buffaloes in Class II. The follicle (FL) area in Class II buffaloes was larger (p < 0.01) than that of the other classes. No influence of the THI on the total PR was recorded. The pregnancy outcome at TAI was affected by the FL area (odds ratio = 2.237; p < 0.05) and body condition score (BCS) (odds ratio = 1.256; p < 0.05). In conclusion, treatment with vaginal P4 optimizes pregnancy rates in anoestrus buffaloes, particularly when the animals are in mid-lactation and show an optimal BCS. Furthermore, the THI does not seem to affect the efficiency of the progesterone treatment

GRADES: Gradient descent for similarity caching

International audienceA similarity cache can reply to a query for an object with similar objects stored locally. In some applications of similarity caches, queries and objects are naturally represented as points in a continuous space. Examples include 360° videos where user's head orientation-expressed in spherical coordinates determines what part of the video needs to be retrieved, and recommendation systems where the objects are embedded in a finite-dimensional space with a distance metric to capture content dissimilarity. Existing similarity caching policies are simple modifications of classic policies like LRU, LFU, and qLRU and ignore the continuous nature of the space where objects are embedded. In this paper, we propose GRADES, a new similarity caching policy that uses gradient descent to navigate the continuous space and find the optimal objects to store in the cache. We provide theoretical convergence guarantees and show GRADES increases the similarity of the objects served by the cache in both applications mentioned above

Relationship among production traits, somatic cell score and temperature–humidity index in the Italian Mediterranean Buffalo

The temperature–humidity index (THI) has been commonly used to analyse heat stress in dairy cattle, but little is known about its effects on buffaloes. In this study, daily milk yield (MY), fat percentage (FP), protein percentage (PP) and somatic cell count (SCC) data from 808 buffalo cows plus environmental temperature and relative humidity were used to investigate the consequence of heat stress. Two mixed models were used to evaluate the impact of THI on MY, FP, PP and log transformed SCC (SCS). The effect of THI was significant for PP, FP and SCS, whereas its interaction with parity was statistically significant for PP and SCS. The relationship between PP and FP and THI was positive but of different magnitude according to the parity. When THI was below 62, an unfavourable effect was observed, especially in primiparous buffalo cows. A significant interaction between SCS and THI across parities was also observed. The effect of THI on MY across parities was not definite but overall a favourable relationship was observed. Our findings depict a susceptibility of buffaloes to low values of THI, suggesting an optimal THI range for water buffaloes between 59 and 63, although some deleterious effects were observed in primiparous buffaloes at THI values lower than 62. Additional investigations are needed to better elucidate the influence of THI on buffalo species.HIGHLIGHTS The overall effect of THI on buffalo diverges from what commonly observed in dairy cattle Cold stress affects milk and udder health in buffaloes The effect of THI on buffaloes’ performance depends on parity, with a larger susceptibility in primiparous than pluriparous buffalo cows Udder health in buffaloes, evaluated using somatic cell count, is also affected by THI

\u3csup\u3e13\u3c/sup\u3eC NMR Investigation of Nonenzymatic Glucosylation of Protein

Nonenzymatic glucosylation of protein is initiated by the reversible condensation of glucose in its open chain form with the amino groups on the protein. The initial product is an aldimine (Schiff base) which cyclizes to the glycosylamine derivative. The aldimine can undergo a slow Amadori rearrangement to yield the relatively stable ketoamine adduct which is structurally analogous to fructose. 13C NMR has been used to characterize these early products of nonenzymatic glucosylation, using RNase A as a model protein. C-1 of the beta-pyranose anomer of the glycosylamine was identified at 88.8 ppm in the spectrum of RNase glucosylated approximately 1:1 with D-[1-13C]glucose. C-1 of the Amadori product was also apparent in this spectrum, resonating as a pair of intense peaks at 52.7 and 53.1 ppm. The anomeric (C-2) resonances of the Amadori adduct were seen in the spectrum of RNase glucosylated approximately 1:1 with [U-13C]glucose. This spectrum was interpreted by comparison to the spectra of reference compounds: D-fructose, fructose-glycine, N alpha-formyl-N epsilon-fructose-lysine, and glucosylated poly-L-lysine. In the protein spectrum, the most intense of the C-2 resonances was that of the beta-fructopyranose anomer at 95.8 ppm. The alpha- and beta-fructofuranose anomers were also observed at 101.7 and 99.2 ppm, respectively. One unidentified signal in the anomeric region was observed in the spectra of poly-L-lysine and RNase, both glucosylated with [U-13C]glucose; no comparable resonances were observed in the spectra of the model compounds

Relationship of body condition score and blood urea and ammonia to pregnancy in Italian Mediterranean buffaloes

The relationship of body condition score ( BCS) and blood urea and ammonia to pregnancy outcome was examined in Italian Mediterranean Buffalo cows mated by AI. The study was conducted on 150 buffaloes at 145 +/- 83 days in milk that were fed a diet comprising 14.8% crude protein, 0.9 milk forage units . kg(-1) dry matter and a non- structural carbohydrate/ crude protein ratio of 2.14. The stage of the oestrous cycle was synchronised by the Ovsynch- TAI programme and blood urea and ammonia levels were assessed on the day of AI. Energy corrected milk ( ECM) production and BCS were recorded bi- weekly. The pregnancy risk was 46.7% and was slightly lower in buffaloes with BCS 7.5. There were no significant differences in ECM, urea and ammonia between pregnant and non- pregnant buffaloes. However, pregnancy outcome was higher ( P = 0.02) in buffaloes with blood urea < 6.83 mmol . L-1. The likelihood of pregnancy for buffaloes with low urea blood level was 2.6 greater than for high urea level and exposure to a high urea level lowered the probability of pregnancy by about 0.25. The findings indicate that buffaloes are similar to cattle and increased blood levels of urea are associated with reduced fertility when animals are mated by AI

Strategies to reduce embryonic mortality in buffalo cows

The aim of the present study was to examine whether treatment with a GnRH agonist, hCG or P4 on Day 25 after AI increased P4 concentrations and reduced the incidence of embryonic mortality (EM) in pregnant buffaloes mated in mid-winter in a Mediterranean environment. The trial was carried out in two farms characterized, in previous years, by low (LEM Group), 153 buffaloes (DIM=150±7 days), and high (HEM Group), 284 buffaloes (DIM=163±5 days), incidence of embryo mortality. Animals were synchronized by Ovsynch-TAI Program and artificially inseminated. On day 25, pregnant buffaloes were randomly assigned to four groups: Control (no treatment), GnRH agonist (buserelin acetate, 12.6 μg), hCG (1500 IU) and P4 (341 mg of P4 i.m. every 4 days for three times). Progesterone (pg/ml) was determined in milk whey on Days 10, 20 and 25 after AI in all buffaloes and in Days 30 and 45 only in buffaloes pregnant on day 25 and assigned to four groups of treatment. Pregnancy diagnosis was undertaken on Day 45 by ultrasound. All treatments increased P4 milk whey and reduced embryonic mortality in buffalo cows bred in the farm characterized by high EM

Buffalo milk and rumen fluid metabolome are significantly affected by green feed

The use of green feed for livestock breeding is an important strategy to encounter both the increasing demand for animal derived products and the perceptions of the consumers regarding animal welfare and sustainability. The aim of this study was to compare different feeding strategies in lactating water buffaloes by using a metabolomic approach. The study was carried out on 32 milking buffaloes that were randomly divided into two groups for a total period of 90 days (3 sampling times). DD Group (dry diet) received a standard total mixed ratio (TMR) characterized by dry forages and concentrates; ZG Group (zero grazing) fed an isoenergetic and isoproteic diet obtained using 30% of sorghum as green forage. Samples of milk and rumen fluid were analyzed by liquid chromatography—mass spectrometry (LC–MS) techniques. Data analyses revealed the presence of several differentially accumulated metabolites and among these, ten compounds were putatively identified in milk samples (i.e. l-carnitine, acetylcarnitine, propionylcarnitine, butyrylcarnitine, 2-methylbutyroylcarnitine, 2-hexenoylcarnitine, hexanoylcarnitine, glycerophosphocholine, δ-valerobetaine and γ-butyrobetaine) and four in rumen fluid (3-(2-hydroxyphenyl) propanoate, Indole-3-acrylic acid, oleamide (cis-9,10-octadecenoamide) and 20-carboxy-leukotriene B4). The modulation of these molecules in buffalo milk is significantly related to the green/dry based feeding and some the natural compound detected could be considered as health-promoting nutrients

Effect of Phosphate on the Kinetics and Specificity of Glycation of Protein

The glycation (nonenzymatic glycosylation) of several proteins was studied in various buffiner os rder to assess the effects of buffering ions on the kinetics and specificity of glycation of protein. Incubation of RNase with glucose in phosphate buffer resulted in inactivation of the enzyme because of preferential modification of lysine residues ino r near the activsei te. In contrast, in the cationic buffers, 3-(N-morpholino)propanesulfonic acid and 3-(N-tris(hydroxymethyl)rnethylamino)- 2-hydroxypropanesulfonica cid, the kineticso f glycation of RNase were decreased 2- to 3-fold, there was a decrease in glycation of active site versus peripheral lysines, and the enzyme was resistant to inactivation by glucose. The extent of Schiff base formation on RNase was comparable in the three buffers, suggesting that phosphate, bound in the active site of RNase, catalyzed the Amadori rearrangement at active site lysines, leading to the enhanced rate of inactivation of the enzyme. Phosphate catalysis of glycation was concentration-dependent and could be mimicked by arsenate. Phosphate also stimulated the rate of glycation of other proteins, such as lysozyme, cytochrome c, albumin, and hemoglobin. As with RNase, phosphate affected the specificity of glycation of hemoglobin, resulting in increasegdly cation of amino-terminal valine versus intrachain lysine residues. 2,3-Diphosphoglycerate exerted similar effeocnt st he glycation of hemoglobin, suggesting that inorganic and organic phosphates may play an important role in determining the kinetics and specificity of glycation of hemoglobin in the red cell. Overall, these studies establishth at buffering ions or ligands can exert significant effects on the kinetics ands pecificity of glycation of proteins