C/C++ Coding for Matrix Pseudo Inverses in Clinical Near Infrared Spectroscopy

Near infrared spectroscopy is used clinically to investigate patterns of change in cerebral oxygenation. We have shown that differences reported between authors are likely the result of computer encoding errors in the manipulation of matrices. Current methods compute the inverse of a non-square matrix to derive chromophore concentration values, and solution of another non-square matrix to derive polynomial coefficients of a least squares best fit curve from which the first derivative can be used to estimate blood flow values. Encoding of these pseudo inverses involves too many nested looping steps to easily identify encoding errors. We have given C/C++ source code along with sample numerical values at the termination of each loop within the algorithm. This provides counter checking for future software development by other programmers, and also permits other investigators to report whether the software used for their experiments agrees with previously published material.     

Amino acid encoding schemes from protein structure alignments: multi-dimensional vectors to describe residue types

Bioinformatic software has used various numerical encoding schemes to describe amino acid sequences. Orthogonal encoding, employing 20 numbers to describe the amino acid type of one protein residue, is often used with artificial neural network (ANN) models. However, this can increase the model complexity, thus leading to difficulty in implementation and poor performance. Here, we use ANNs to derive encoding schemes for the amino acid types from protein three-dimensional structure alignments. Each of the 20 amino acid types is characterized with a few real numbers. Our schemes are tested on the simulation of amino acid substitution matrices. These simplified schemes outperform the orthogonal encoding on small data sets. Using one of these encoding schemes, we generate a colouring scheme for the amino acids in which comparable amino acids are in similar colours. We expect it to be useful for visual inspection and manual editing of protein multiple sequence alignments.     

Complement component C8gamma is expressed in human fetal and adult kidney independent of C8alpha

Human complement component C8gamma is an unusual complement factor since it shows no homology to other complement proteins but is a member of the lipocalin superfamily. So far, it has been found exclusively in plasma, covalently linked to C8alpha by disulfide bridging. We have used dot blot and Northern blot analyses of a large number of different human tissues to survey systematically the expression pattern of C8gamma. Our experiments clearly showed that besides in liver, this gene is also expressed in fetal and adult kidney. Renal expression of C8gamma is not dependent on C8alpha expression, since we could not detect C8alpha expression in kidney. Thus its physiological function is not restricted to a specific action in association with complement components. As a prerequisite for further characterization of the structure and binding activities of the uncomplexed C8gamma, we have expressed the encoding cDNA in Escherichia coli. To increase the probability for proper folding of the characteristic intramolecular disulfide bridge the recombinant protein was produced by secretion to the periplasm.     

Fisher information matrix of the Dirichlet-multinomial distribution

In this paper we derive explicit expressions for the elements of the exact Fisher information matrix of the Dirichlet-multinomial distribution. We show that exact calculation is based on the beta-binomial probability function rather than that of the Dirichlet-multinomial and this makes the exact calculation quite easy. The exact results are expected to be useful for the calculation of standard errors of the maximum likelihood estimates of the beta-binomial parameters and those of the Dirichlet-multinomial parameters for data that arise in practice in toxicology and other similar fields. Standard errors of the maximum likelihood estimates of the beta-binomial parameters and those of the Dirichlet-multinomial parameters, based on the exact and the asymptotic Fisher information matrix based on the Dirichlet distribution, are obtained for a set of data from Haseman and Soares (1976), a dataset from Mosimann (1962) and a more recent dataset from Chen, Kodell, Howe and Gaylor (1991). There is substantial difference between the standard errors of the estimates based on the exact Fisher information matrix and those based on the asymptotic Fisher information matrix.     

Kinetic isotope effects significantly influence intracellular metabolite 13C labeling patterns and flux determination

Rigorous mathematical modeling of carbon-labeling experiments allows estimation of fluxes through the pathways of central carbon metabolism, yielding powerful information for basic scientific studies as well as for a wide range of applications. However, the mathematical models that have been developed for flux determination from ¹³C labeling data have commonly neglected the influence of kinetic isotope effects on the distribution of ¹³C label in intracellular metabolites, as these effects have often been assumed to be inconsequential. We have used measurements of the ¹³C isotope effects on the pyruvate dehydrogenase enzyme from the literature to model isotopic fractionation at the pyruvate node and quantify the modeling errors expected to result from the assumption that isotope effects are negligible. We show that under some conditions kinetic isotope effects have a significant impact on the ¹³C labeling patterns of intracellular metabolites, and the errors associated with neglecting isotope effects in ¹³C-metabolic flux analysis models can be comparable in size to measurement errors associated with GC–MS. Thus, kinetic isotope effects must be considered in any rigorous assessment of errors in ¹³C labeling data, goodness-of-fit between model and data, confidence intervals of estimated metabolic fluxes, and statistical significance of differences between estimated metabolic flux distributions.     

Quantitative binding models for CYP2C9 based on benzbromarone analogues

The cytochrome P450 (CYP) isoforms involved in xenobiotic metabolism are enzymes whose substrate selectivity remains difficult to predict due to wide specificity and dynamic protein-substrate interactions. To uncover the determinants of specificity for cytochrome CYP2C9, a novel library of benzbromarone (bzbr) inhibitors was used to reevaluate its pharmacophore. CoMSIA was used with the bzbr ligands to generate both quantitative binding models and three-dimensional contour plots that pinpoint predicted interactions that are important for binding to 2C9. Since this class of compounds is more potent than any other toward 2C9, the small molecule properties deemed most ideal by the software were used to address protein-ligand interactions using new mutagenesis and structural data. Nine new bzbr analogues provide evidence that specific electrostatic and hydrophobic interactions contribute the most to 2C9's specificity. Three of the new analogues are better isosteres of bzbr that contain bulky groups adjacent to the phenol and have increased pK(a) values. These ligands test the hypothesis that anionic substrates bind with higher affinity to 2C9. Since they have higher affinity than the previous nonacidic analogues, the importance of bulky groups on the phenol ring appears to have been underestimated. CoMSIA models predict that these bulky groups are favorable for their hydrophobicity, while a negative charge is favored at the ketone oxygen rather than the phenol oxygen. The overlap of this ketone with electronegative groups of other 2C9 substrates suggests they act as key positive charge acceptors.     

Tracking and Quantifying Developmental Processes in C. elegans Using Open-source Tools

Quantitatively capturing developmental processes is crucial to derive mechanistic models and key to identify and describe mutant phenotypes. Here protocols are presented for preparing embryos and adult C. elegans animals for short- and long-term time-lapse microscopy and methods for tracking and quantification of developmental processes. The methods presented are all based on C. elegans strains available from the Caenorhabditis Genetics Center and on open-source software that can be easily implemented in any laboratory independently of the microscopy system used. A reconstruction of a 3D cell-shape model using the modelling software IMOD, manual tracking of fluorescently-labeled subcellular structures using the multi-purpose image analysis program Endrov, and an analysis of cortical contractile flow using PIVlab (Time-Resolved Digital Particle Image Velocimetry Tool for MATLAB) are shown. It is discussed how these methods can also be deployed to quantitatively capture other developmental processes in different models, e.g., cell tracking and lineage tracing, tracking of vesicle flow.     

The rDNA of C. elegans: sequence and structure.

We have sequenced one complete rDNA tandem repeat from the nematode C. elegans. By comparative analysis we derive secondary structures for the 18s, 5.8s, and 26s rRNA molecules, and comment on other important features of the sequence. We also present the sequence of a junction between the rDNA and non-ribosomal DNA. Finally, we use our data to quantify the evolutionary relationships among several organisms currently studied in developmental biology.     

A detailed kinetic study of Mox-1, a plasmid-encoded class C beta-lactamase

Surveys of beta-lactamases in different parts of the world show an important increase in class C beta-lactamases, thus the study of these enzymes is becoming an important issue. We created an overproduction system for Mox-1, a plasmid class C beta-lactamase, by cloning the gene encoding this enzyme, and placing it under the control of a T7 promoter, using vector pET 28a. The enzyme, purified by ion exchange chromatography, was used to obtain the molecular mass (38246), the N-terminal sequence (GEASPVDPLRPVV), and pI (8.9), and to perform a detailed kinetic study. Cephalotin was used as reporter substrate in the case of poor substrates. The kinetic study showed that benzylpenicillin, cephalotin, cefcapene and moxalactam were good substrates for Mox-1 (k(cat)/K(m) values >2.5 x 10(6) M(-1) s(-1)). On the other hand, ceftazidime and cefepime were poor substrates for this enzyme (K(m) values >200 microM). Clavulanic acid had no inhibitory effect on Mox-1 (K(m)=30.2 mM), however aztreonam behaved as an inhibitor of Mox-1 (K(i)=2.85 microM).     

Minimizing errors in the analysis of dive recordings from shallow-diving animals

Knowledge of the diving behaviour of aquatic animals expanded considerably with the invention of time-depth recorders (TDRs) in the 1960s. The large volume of data acquired from TDRs can be analyzed using dive analysis software, however, the application of the software has received relatively little attention. We present an empirical procedure to select optimum values that are critical to obtaining reliable results: the zero-offset correction (ZOC) and the dive threshold. We used dive data from shallow-diving coastal dugongs (Dugong dugon) and visual observations from an independent study to develop and test a procedure that minimizes errors in characterizing dives. We initially corrected the surface level using custom software. We then determined the optimum values for each parameter by classifying dives identified by an open-source dive analysis software into Plausible and Implausible dives based on the duration of dives. The Plausible dives were further classified as Unrecognized dives if they were not identified by the software but were of realistic dive duration. The comparison of these dive types indicated that a ZOC of 1 m and a dive threshold of 0.75 m were the optimum values for our dugong data as they gave the largest number of Plausible dives and smaller numbers of other dive types. Frequency distributions of dive durations from TDRs and independent visual observations supported the selection. Our procedure could be applied to other shallow-diving animals such as coastal dolphins and turtles.     

Effects of temperature and composition on the viscosity of respiratory gases

The steady-state sensitivity of resistance pneumotachographs is proportional to viscosity. Dynamic characteristics of pneumotachographs, pressure transducers, and mass spectrometers are also viscosity dependent. We derive linear equations to approximate the viscosities of O2, N2, CO2, H2O, He, N2O, and Ar for temperatures between 20 and 40 degrees C by using published viscosity data and a nonlinear extrapolation equation. We verify the accuracy of the extrapolation equation by comparison with published data. Our linear equations for pure gas viscosities yield standard errors less than 0.35 microP. We also compare a nonlinear equation for calculating the viscosities of mixtures of gases with published measured viscosities of dry air, humid air, and He-O2 and N2-CO2 mixtures. The maximum difference between published and calculated values is 1.3% for 10% CO2 in N2. All other differences are less than 0.38%. For saturated humid air at 35 degrees C, a linear concentration-weighted combination of viscosities differs from our nonlinear equation by 4.9, 2.1, and 1.7% at barometric pressures of 32, 83, and 100 kPa, respectively. By use of our method, the viscosity of normal respiratory gases can be calculated to within 1% of measured values.     

Measuring and comparing evolvability and constraint in multivariate characters

The Lande equation forms the basis for our understanding of the short-term evolution of quantitative traits in a multivariate context. It predicts the response to selection as the product of an additive genetic variance matrix and a selection gradient. The selection gradient approximates the force and direction of selection, and the genetic variance matrix quantifies the role of the genetic system in evolution. Attempts to understand the evolutionary significance of the genetic variance matrix are hampered by the fact that the majority of the methods used to characterize and compare variance matrices have not been derived in an explicit theoretical context. We use the Lande equation to derive new measures of the ability of a variance matrix to allow or constrain evolution in any direction in phenotype space. Evolvability captures the ability of a population to evolve in the direction of selection when stabilizing selection is absent. Conditional evolvability captures the ability of a population to respond to directional selection in the presence of stabilizing selection on other trait combinations. We then derive measures of character autonomy and integration from these evolvabilities. We study the properties of these measures and show how they can be used to interpret and compare variance matrices. As an illustration, we show that divergence of wing shape in the dipteran family Drosophilidae has proceeded in directions that have relatively high evolvabilities.     

The receptor for the subgroup C avian sarcoma and leukosis viruses, Tvc, is related to mammalian butyrophilins, members of the immunoglobulin superfamily

The five highly related envelope subgroups of the avian sarcoma and leukosis viruses (ASLVs), subgroup A [ASLV(A)] to ASLV(E), are thought to have evolved from an ancestral envelope glycoprotein yet utilize different cellular proteins as receptors. Alleles encoding the subgroup A ASLV receptors (Tva), members of the low-density lipoprotein receptor family, and the subgroup B, D, and E ASLV receptors (Tvb), members of the tumor necrosis factor receptor family, have been identified and cloned. However, alleles encoding the subgroup C ASLV receptors (Tvc) have not been cloned. Previously, we established a genetic linkage between tvc and several other nearby genetic markers on chicken chromosome 28, including tva. In this study, we used this information to clone the tvc gene and identify the Tvc receptor. A bacterial artificial chromosome containing a portion of chicken chromosome 28 that conferred susceptibility to ASLV(C) infection was identified. The tvc gene was identified on this genomic DNA fragment and encodes a 488-amino-acid protein most closely related to mammalian butyrophilins, members of the immunoglobulin protein family. We subsequently cloned cDNAs encoding Tvc that confer susceptibility to infection by subgroup C viruses in chicken cells resistant to ASLV(C) infection and in mammalian cells that do not normally express functional ASLV receptors. In addition, normally susceptible chicken DT40 cells were resistant to ASLV(C) infection after both tvc alleles were disrupted by homologous recombination. Tvc binds the ASLV(C) envelope glycoproteins with low-nanomolar affinity, an affinity similar to that of binding of Tva and Tvb with their respective envelope glycoproteins. We have also identified a mutation in the tvc gene in line L15 chickens that explains why this line is resistant to ASLV(C) infection.     

H1° and H3.3B mRNA levels in developing rat brain

Two overlapping rat cDNAs, covering a continuous region of 1107 base pairs, have been isolated and sequenced. The clones contain identical open reading frames, encoding a 136 amino acid long polypeptide which exhibits 100% identity to other mammalian H3.3 histone variants. We show that the inserts derive, in particular, from the H3.3B gene. We used these inserts and an insert from an H1° encoding clone, previously described (6), as probes to study the accumulation of mRNAs encoding the corresponding histone replacement variants (namely, H1° and H3.3) during rat brain development. We found that the concentration of both H1° and H3.3B mRNAs decreases from the embryonal day 18 (E18) to the postnatal day 10 (P10), with inverse correlation to protein accumulation.This paper is dedicated to our friend Paolo Carbone who devoted his life to research and teaching in Genetics. We will always remember him for scientific honesty and for his unique qualities of humanity.     

The use of various properties of amino acids in color and monochrome dot-matrix analyses for protein homologies

Software has been developed to allow the use of a number ofparameters in the comparative representation of proteins incolor and monochrome dot matrices. They include the parametersof partial specific volume, residue bulkiness, the mean areaburied of side chains, seven additional hydropathy scales, mutability,polarity, secondary structure propensities, energy/residue,energy/atom, R_f values, the pKs at the N and C terminals, user-definedparameters and, if desired, randomly generated values. Manyof these parameters can be combined in n space using an algorithmbased on the Euclidian distance relationship in order to deriveconsensus values. The problem of scoring matched identitiesis addressed and the user may stipulate that they score 100on a 0–100 scale or be determined from the Dayhoff MDM₇₈values with the rest of the matrix scaled appropriately. ThePAMs matrix has been incorporated in such a way to allow theuser to stipulate various PAM's values or estimated percentagedifference between two peptide sequences, and converting tolog odds values. In addition, the similarity ring developedby Swanson and the matrix proposed by Bacon and Anderson havebeen adapted for use in the program. Color indices have beenutilized to give a ‘third dimension’ to the projections,allowing the user to judge the degree of similarity of differentregions which are represented. The software also provides forthe plotting of nucleotides in which case color is used to codeindividual nucleotides, purines versus pyrimidines, or similarcolors are used to differentiate between A and T bases on theone hand, and G and C on the other. Received on December 31, 1987; accepted on May 18, 1988     

Bioinformatic identification of genes encoding C1q-domain containing proteins in zebrafish

C1q is the first subcomponent of classical pathway in the complement system and a major link between innate and acquired immunities. The globular (gC1q) domain similar with C1q was also found in many non-complement C1q-domain-containing (C1qDC) proteins which have similar crystal structure to that of the multifunctional tumor necrosis factor (TNF) ligand family, and also have diverse functions. In this study, we identified a total of 52 independent gene sequences encoding C1q-domain-containing proteins through comprehensive searches of zebrafish genome, cDNA and EST databases. In comparison to 31 orthologous genes in human and different numbers in other species, a significant selective pressure was suggested during vertebrate evolution. Domain organization of C1q-domain-containing (C1qDC) proteins mainly includes a leading signal peptide, a collagen-like region of variable length, and a C-terminal C1q domain. There are 11 highly conserved residues within the C1q domain, among which 2 are invariant within the zebrafish gene set. A more extensive database searches also revealed homologous C1qDC proteins in other vertebrates, invertebrates and even bacterium, but no homologous sequences for encoding C1qDC proteins were found in many species that have a more recent evolutionary history with zebrafish. Therefore, further studies on C1q-domain-containing genes among different species will help us understand evolutionary mechanism of innate and acquired immunities.     

MUCIDS: an operative C environment for acquisition and processing of polarized-light scattered from biological specimens

In this work, we describe a software package, MUCIDS, completelydeveloped in our laboratory, for acquisition and processingof differential polarizxition light-scattering data from specimensof biophysical interest. MUCIDS is a C environment that managesthe whole activity of an instrument used for measurements ofMueller matrix scattering elements. It allows one to capture,analyse, process and display data from this or from other similarlight-scattering experiments. The entire system is suitablefor routine measurements in a general biophysical (or microbiological)laboratory because of its easy handling and maintenance. Thesoftware was written in C lattice and will run on IBM personalcomputers and similar. It uses IBM/DAC and GPIB/IBM interfacecards. Received on October 3, 1989; accepted on March 26, 1990     

Three-compartment model: critical evaluation based on neutron activation analysis

There is renewed interest in Siri's classic three-compartment (3C) body composition model, requiring body volume (BV) and total body water (TBW) estimates, because dual-energy X-ray absorptiometry (DEXA) and in vivo neutron activation (IVNA) systems cannot accommodate subjects with severe obesity. However, the 3C model assumption of a constant ratio (alpha) of mineral (M) to total body protein (TBPro) and related residual mass density (D(RES)) based on cadaver analyses might not be valid across groups differing in sex, race, age, and weight. The aim of this study was to derive new 3C model coefficients in vivo and to compare these estimates to those derived by Siri. Healthy adults (n = 323) were evaluated with IVNA and DEXA and the measured components used to derive alpha and D(RES). For all subjects combined, values of alpha and D(RES) (means +/- SD, 0.351 +/- 0.043; 1.565 +/- 0.023 kg/l) were similar to Siri's proposed values of 0.35 and 1.565 kg/l, respectively. However, alpha and D(RES) varied significantly as a function of sex, race, weight, and age. Expected errors in percent body fat arising by application of Siri's model were illustrated in a second group of 264 adults, including some whose size exceeded DEXA limits but whose BV and TBW had been measured by hydrodensitometry and (2)H(2)O dilution, respectively. Extrapolation of predictions by newly developed models to very high weights allows percent fat error estimation when Siri's model is applied in morbidly obese subjects. The present study results provide a critical evaluation of potential errors in the classic 3C model and present new formulas for use in selected populations.