Analysis of ligand-binding data without knowledge of bound or free ligand molar concentration

A method is proposed to set the parameters in a nonlinear regression procedure to determine the equilibrium dissociation constant (Kd) and the high affinity receptor concentration (Bmax) of systems consisting of one ligand, one high affinity receptor, and n low affinity binding sites. This method is suitable when neither bound or free ligand formal concentrations nor the maximum of the binding signal can be deduced from the experimental data. The method makes use of (i) the abscissa of the first inflection point of the plot of any signal proportional to the binding of ligand to receptors versus the logarithm of the total ligand concentration, and (ii) the initial slope of the saturation curve plotted in direct coordinates. We first demonstrate that when such an inflection point exists, its abscissa lies between Bmax/2 + Kd(1 + d) and Bmax + 2Kd(1 + d), where d is a parameter representative of the binding to the low affinity sites. Second, we demonstrate that the initial slopes of two saturation curves in direct coordinates, where Bmax varies by a known factor, allows an estimation of the Bmax/Kd ratio, within certain limits. From these two sets of data it is subsequently possible to define a precise window for the values of both Bmax and Kd. The performance of the method has been evaluated in representative cases using Monte Carlo studies. The results establish conditions for the existence of an inflection point as well as the influence of low affinity binding, whether or not proportional to Bmax.     

Formation of glyceryl 2-mononitrate by regioselective bioconversion of glyceryl trinitrate: efficiency of the filamentous fungus Phanerochaete chrysosporium

Various microorganisms have been evaluated for their ability to hydrolyze glyceryl trinitrate (GTN) to glyceryl dinitrates and mononitrates. Provided that the GTN extracellular concentration was under the lethal dose, metabolite formation and regioselectivity depend on the nature of the strain used. In particular, Phanerochaete chrysosporium at a sublethal dose (3 mM) converts GTN into 1,2-glyceryl dinitrate and 2-glyceryl mononitrate (2-GMN) with a 80% regioselectivity in both steps. This bioconversion, when carried out in fermentors at 28 degrees C, allowed formation of 2-GMN at a rate of 12 mumol/h/g of dried mycelium. Successive batches of 3 mM GTN could be converted into 2-GMN as long as consecutive additions of glycerol or glucose were effected to ensure cell survival and the efficiency of the enzymatic system involved.     

Nucleotide sequence evidence of the unicentric origin of the βC mutation in Africa

Summary The origin of the ^C mutation was studied by characterizing nucleotide sequence polymorphisms on ^C chromosomes of patients from various African countries. In the majority of cases, the ^C mutation was found in linkage disequilibrium with a single chromosomal structure as defined by classical RFLP haplotypes, intergenic nucleotide sequence polymorphisms immediately upstream of the -globin gene, and intragenic -globin gene polymorphisms (frameworks). In addition, three atypical variant chromosomes carrying the ^C mutation were observed, and are most probably explained either by a meiotic recombination (two cases) or by one nucleotide substitution occurring in an unstable array of tandemly repeated sequences (one case). These data demonstrate the unicentric origin of the ^C mutation in central West Africa, with subsequent mutational modification in a small number of instances. The data also supports gene flow of the ^C chromosome from subsaharan Africa to North Africa.     

The stereochemistry of hydrogen abstraction in vitamin K-dependent carboxylation

The stereochemistry of the hydrogen abstraction in the vitamin K-dependent carboxylation of synthetic peptides has been investigated; the carboxylation rates of various peptidic substrates containing a stereospecifically 4-monodeuterated glutamic acid residue have been compared to that of nondeuterated peptides. A significant isotope effect was found only with the substrates containing (4S)-4-deuterated glutamic acid. These data reveal that the rat liver microsomal vitamin K-dependent carboxylase acts stereospecifically in abstracting the 4-pro-S hydrogen of the glutamyl residue. The low values of the measured isotope effects indicate that the hydrogen abstraction does not constitute a limiting step in the carboxylation mechanism.     

A stochastic simulation study on using different models for prediction of breeding values while changing the breeding goal

In a stochastic simulation study the effect of simultaneously changing the model for prediction of breeding values and changing the breeding goal was studied. A population of 100 000 cows with registrations on seven traits was simulated in two steps. In the first step of 15 years the population was selected for production and mastitis occurrence using a univariate model for prediction of breeding values for production and a trivariate model using information on mastitis treatments, udder depth and somatic cell score for prediction of breeding values for mastitis occurrence. In the second step six different scenarios were set up and simulated for 15 years combining two different breeding goals and three different models for prediction of breeding values in 20 replicates. Breeding goal 1 had relative economic value per genetic standard deviation on production (19.4) and mastitis occurrence ( − 50) whereas breeding goal 2 had a economic value on production (19.4), udder depth (4.2), mastitis occurrence ( − 50), non return rate (13.0) and days open ( − 16.75). Model 1 was a model similar to the one used in the first 15 years. Model 2 was an approximate multitrait model where solutions for fixed effects from a model corresponding to model 1 were subtracted from the phenotypes and a multitrait model with an overall mean, a year effect, an additive genetic and a residual effect were applied. Model 3 was a full multitrait model. Average genetic trends for total merit and each individual trait over 20 replicates were compared for each scenario. With the number of replicates the genetic responses using model 2 and 3 were not significant different. With a broad breeding goal using, model 2 or model 3 gave a significantly higher response in total merit than using model 1. Using a narrow breeding goal there was no significant difference between models used for prediction of breeding values. Results showed that with a breeding goal with a lot of emphasis on low heritable traits with a high economic value using a multitrait methodology for prediction of breeding values will redistribute the genetic progress in the total merit index. More gain will come from the low heritable traits in the breeding goal and less from traits with higher heritability. With a broad breeding goal and exploiting the available information in the data the inbreeding coefficient increased though not significantly.     

Modelling and estimation of genotype by environment interactions for production traits in French dairy cattle

Background

Genotype by environment interactions are currently ignored in national genetic evaluations of dairy cattle. However, this is often questioned, especially when environment or herd management is wide-ranging. The aim of this study was to assess genotype by environment interactions for production traits (milk, protein, fat yields and fat and protein contents) in French dairy cattle using an original approach to characterize the environments.

Methods

Genetic parameters of production traits were estimated for three breeds (Holstein, Normande and Montbéliarde) using multiple-trait and reaction norm models. Variables derived from Herd Test Day profiles obtained after a test day model evaluation were used to define herd environment.

Results

Multiple-trait and reaction norm models gave similar results. Genetic correlations were very close to unity for all traits, except between some extreme environments. However, a relatively wide range of heritabilities by trait and breed was found across environments. This was more the case for milk, protein and fat yields than for protein and fat contents.

Conclusions

No real reranking of animals was observed across environments. However, a significant scale effect exists: the more intensive the herd management for milk yield, the larger the heritability.     

Binational evaluation of type traits from Germany and France with a single-trait MACE animal model

Binational genetic evaluation between Germany and France were performed for each type trait using a single-trait MACE (multiple across-country evaluation) model. Daughter yield deviations (DYD) of bulls having 30 equivalent daughter contributions or more were the data for parameter estimation. Full pedigree information of bulls was used via sire and dam relationships. In general, across-country genetic correlation estimates were in agreement with what is observed by Interbull. The estimated correlations were over 0.93 for stature, rump angle, udder depth, front teat placement, teat length and rear teat placement. These traits have been classified in both countries for a long period of time. However, some other type traits were included later in the French type classification system (most of them since 2000): chest width, body depth, angularity, rump width, rear leg rear view, fore udder and rear udder height. The estimated correlations for these traits were relatively low. In order to check changes in genetic correlations over time, data from bulls born until the end of 1995 were discarded. Higher genetic correlation estimates between both countries were obtained by using more recent data especially for traits having lower genetic correlation, e.g. body depth correlation increased from 0.55 to 0.83. Once genetic correlations were estimated, binational genetic evaluation between Germany and France were performed for each type trait using DYD of bulls. The rankings of bulls obtained from this evaluation had some differences with Interbull rankings but a similar proportion of bulls from each country was found. Finally, more computationally demanding binational evaluations were performed using yield deviations of cows for binational cow comparison. The rankings obtained were influenced by the number of daughters per bull and heritabilities used in each country.     

Validation of an approximate approach to compute genetic correlations between longevity and linear traits

The estimation of genetic correlations between a nonlinear trait such as longevity and linear traits is computationally difficult on large datasets. A two-step approach was proposed and was checked via simulation. First, univariate analyses were performed to get genetic variance estimates and to compute pseudo-records and their associated weights. These pseudo-records were virtual performances free of all environmental effects that can be used in a BLUP animal model, leading to the same breeding values as in the (possibly nonlinear) initial analyses. By combining these pseudo-records in a multiple trait model and fixing the genetic and residual variances to their values computed during the first step, we obtained correlation estimates by AI-REML and approximate MT-BLUP predicted breeding values that blend direct and indirect information on longevity. Mean genetic correlations and reliabilities obtained on simulated data confirmed the suitability of this approach in a wide range of situations. When nonzero residual correlations exist between traits, a sire model gave nearly unbiased estimates of genetic correlations, while the animal model estimates were biased upwards. Finally, when an incorrect genetic trend was simulated to lead to biased pseudo-records, a joint analysis including a time effect could adequately correct for this bias.