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Anders Fuglsang

Publications and source records attributed to Anders Fuglsang.

At least 19 recordsLinked to original sources
PubMed2006

Accounting for background nucleotide composition when measuring codon usage bias: brilliant idea, difficult in practice.

The effective number of codons used in a gene is a commonly used measure of codon usage. It varies between 20 and 61 (standard genetic code) and indicates to which degree the entire genetic code is used. It is a drawback of this method that it does not take background composition into account. This led Novembre to introduce a variant called Nc' (Novembre JA. 2002. Accounting for background nucleotide composition when measuring codon usage bias. Mol Biol Evol 19:1390-4). In this letter, its properties are under the loupe, with special emphasis on phenomena relating to codon homozygosity. A theoretical misunderstanding regarding this estimator is explained in detail, notably Nc varies between 0 and 61 instead of 20 and 61 (with the standard genetic code). Practical examples from the genome of Pseudomonas aeruginosa are given which demonstrate that the problem is not just theoretical.

Base Composition↗
PubMed2005

Estimating the "effective number of codons": the Wright way of determining codon homozygosity leads to superior estimates.

In 1990, Frank Wright introduced a method for measuring synonymous codon usage bias in a gene by estimation of the "effective number of codons," N(c). Several attempts have been made recently to improve Wright's estimate of N(c), but the methods that work in cases where a gene encodes a protein not containing all amino acids with degenerate codons have not been tested against each other. In this article I derive five new estimators of N(c) and test them together with the two published estimators, using resampling under rigorous testing conditions. Estimation of codon homozygosity, F, turns out to be a key to the estimation of N(c). F can be estimated in two closely related ways, corresponding to sampling with or without replacement, the latter being what Wright used. The N(c) methods that are based on sampling without replacement showed much better accuracy at short gene lengths than those based on sampling with replacement, indicating that Wright's homozygosity method is superior. Surprisingly, the methods based on sampling with replacement displayed a superior correlation with mRNA levels in Escherichia coli.

Codon↗
PubMed2005

Analysis of 5' UTR composition and gene expression: canonical versus non-canonical start codons.

The overall composition upstream of start codons in Escherichia coli was evaluated and viewed in connection with global transcriptome data. Genes starting with AUG as initiation codon tended to be expressed at higher levels than the non-AUG genes, and the upstream region of the non-AUG genes showed negligible signs of Shine-Dalgarno sequences. The latter is in sharp contrast to the AUG genes. Viewing these findings in connection with the current literature, it is proposed that a distinct mechanism for initiation of translation might exist for non-AUG genes that are not preceded by a Shine-Dalgarno sequence. A survey covering a range of other eubacteria (Firmicutes, Proteoacteria, and Actinobacteria) reveals that it is mainly among the Proteobacteria that non-AUG genes do not display clear signs of Shine-Dalgarno regions.

5' Untranslated Regions↗
PubMed2005

On the methodological weakness of 'the effective number of codons': a reply to Marashi and Najafabadi.

In a recent publication [Biochem. Biophys. Res. Commun. 317 (2004) 957] it was proposed that the 'effective number of codons' (Nc) in a gene should be calculated by summing the individual amino acid Nc's using rounding whenever the codon homozygosities are lower than the reciprocal value of the number of members of the synonymous families. This led Marashi and Najafabadi to examine the consequences of individual re-adjustment when comparing observed Nc with the expected Nc under assumptions of no selection, and C=G and A=T [Biochem. Biophys. Res. Commun. 324 (2004) 1]. Clearly, the present methodology has some weaknesses; in this work, I discuss these in relation to the observations by Marashi and Najafabadi, and finally an alternative method for the calculation of Nc is introduced with the purpose of eliminating the need for re-adjustments.

Bias↗
PubMed2005

Correlation of codon bias measures with mRNA levels: analysis of transcriptome data from Escherichia coli.

Although codon usage is often represented by a 61-dimensional vector, the ability of determining the codon bias in a gene relies on a uni-dimensional vector which measures the total bias in usage of synonymous codons. Codon usage is receiving more and more focus because codon biases might be valuable tools to predict and optimize gene/protein expression. How good any of these measures is for correlating codon usage with gene and protein expression has yet to be investigated. In this study, we correlated gene transcript levels in Escherichia coli with codon usage, using a number of different codon bias measures. We found that there is a significant correlation between transcript levels and codon bias measures, suggesting that these measures can be used to assess or predict gene expression. The codon bias measure performing best in this context was the codon adaptation index.

Codon↗
PubMed2005

A cytosine-rich region upstream of start codons serving as a signal for initiation of translation in Encephalitozoon cuniculi?

Encephalitozoon cuniculi, an intracellular eukaryote frequently infecting immunodeficient humans, displays pronounced compaction in its genes. Short-leaded mRNA has been observed which has led to speculation into alternative mechanisms of translation initiation. It has been proposed that a 'downstream box' could serve as an initiation signal. In this report, non-randomness analysis was used to study the genes of E. cuniculi. Surprisingly, it was found that the region 5-10nt upstream of start codons is highly cytosine-enriched and this phenomenon clearly differentiates genes of predicted high and low expressivity. These two groups of genes can, on the other hand, not be clearly differentiated from non-randomness plots downstream of their start codons. The data thus do not support the 'downstream box' hypothesis but seem to suggest that initiation is depending on the region immediately upstream of start codons. The cytosine richness has no known parallel in other eukaryotes, prokaryotes or archaebacteria.

Animals↗
PubMed2004

INTROnSPECTIVE and NORA: computerized compositional characterization of regions within introns and around start codons on the basis of non-randomness analysis.

Non-randomness analysis has recently proven a valuable method in the characterization of chromosomes and genomes with respect to the nucleotides around start codons. This methodology has been implemented in a distributable program NORA, which is presented as freeware for users in academia. Moreover, because the current knowledge about introns is limited, another application, INTROnSPECTIVE, has been developed. This application analyzes introns in either direction (from 5' to 3' or from 3' to 5') and the user has the option to exclude introns on the basis of their size or intron number within the gene. Both programs are based on initial parsing of GenBank flatfiles, i.e., with these programs, entire genomes or chromosomes can be parsed and characterized within seconds. The programs run under 32-bit Windows operating systems and can be obtained via . They are the only ones available which perform non-randomness analysis, and INTROnSPECTIVE represents a comprehensive and novel methodology for the study of introns.

Algorithms↗
PubMed2004

Compositional nonrandomness upstream of start codons in archaebacteria.

Since the regions directing transcriptional and translational initiation in archaebacteria are poorly characterized, the purpose of this study was to characterize them using measurements of nonrandomness upstream of start codons on eight fully sequenced archaebacterial genomes. Two distinctly different regions with conservation were identified. The location of the first corresponded well to the classical Shine-Dalgarno region (phi peak), and the other was located approximately 20-35 nucleotides upstream of start codons (alpha region), but both regions are not present in all strains. The composition of the region around the phi peak showed an overrepresentation of guanine, whereas composition in the alpha region had an overrepresentation of adenine and thymine. It is furthermore shown that the alpha region surprisingly is associated with start codon usage and other characteristics of the genes. The alpha region is likely to correspond to TATA-boxes and thereby indicates use of leaderless (or short-leadered) transcripts.

5' Flanking Region↗
PubMed2004

The 'effective number of codons' revisited.

Frank Wright [Gene 87 (1990) 23] derived a formula for calculation of a quantity termed the 'effective number of codons' (Nc) based on codon homozygosities. This quantity is a number between 20 and 61 and tells to what degree the codon usage in a gene is biased, i.e., it approaches 20 codons for the extremely biased genes, and approaches 61 for the genes where all possible codons are used with no preference. Among the different measures of codon bias Nc is considered the most useful and has found widespread use in papers dealing with codon usage phenomena. In this paper, the mathematical behaviours of codon homozygosities and Nc are evaluated, using Escherichia coli as the model organism. The results indicate that the classical formula for calculation of Nc could appropriately be substituted under circumstances, where there is bias discrepancy, i.e., when one amino acid (or more) within a degeneracy group is associated with strong codon bias while at the same time others in the same degeneracy group have little bias. An alternative estimator, termed Nc, is proposed and tested against Nc, and performs better when there is such bias discrepancy.

Codon↗
PubMed2004

The relationship between palindrome avoidance and intragenic codon usage variations: a Monte Carlo study.

Several studies have shown that codon usage within genes varies, as it seems dependent on both codon context and codon position within the gene. Given that palindromes in addition often are avoided in genomes, this study aimed at finding out if intragenic variations in codon usage may be a way to control the amount and location of palindromes. A Monte Carlo algorithm was written which resampled the codons in genes while keeping the amino acid sequence of the translation product constant. On the resampled sequences, palindromes were counted and their intragenic positions mapped. Escherichia coli K12 uses type II restriction-modification systems and displays pronounced codon usage phenomena. Using this as a reference organism it was clearly shown that the number of palindromes in genes is generally lower than the amount of palindromes in resampled genes; thus, the succession of codons seems to be a way to decrease the number of palindromes. The intragenic position of palindromes in resampled sequences, however, was largely equal to the position in the native genes, so codon usage phenomena are unlikely to be a way to control the intragenic position of palindromes. The analysis was repeated on two bacteriophages and gave similar same results, even though the virus genomes are much smaller. Studies on the endosymbionts Buchnera sp. APS and Wigglesworthia sp., which seemingly have no type II restriction-modification systems, showed that in these species there is only weak evidence for codon usage acting to control the number of palindromes.

Algorithms↗
PubMed2004

Evolution of prokaryotic DNA: intragenic and extragenic divergences observed with orthologs from three related species.

This study compared orthologous gene pairs from Escherichia coli K12, E. coli O157:H7 EDL933, Salmonella typhimurium LT2, and Yersinia pestis CO92 using only homologs of equal length, and differing nucleotides were counted and mapped. The data showed very clearly how the rates of divergence change with intragenic and extragenic position. The rate of synonymous mutation is lowest near start codons and near stop codons, and, a little surprisingly, the opposite seemed to be true for nonsynonymous substitutions. Analysis outside genes reveals that nucleotide divergences occur less frequently upstream of start codons than downstream of stop codons, and a remarkable drop in divergences is seen for two of the data sets around N = 9 nucleotides upstream of start codons; that is, the Shine-Dalgarno region changes at a lower rate. The explanation is likely to be the link between expressivity and sequence complementarity to the 3' end of 16S ribosomal rRNA. The latter is highly conserved across many bacterial and archaebacterial species.

Bacteria↗
PubMed2004

Using spectrophotometry to determine in vitro turnover rates of peptides in plasma.

A new method for determination of first-order elimination constants for dipeptides is presented. The peptides are hydrolysed by plasma enzymes into amino acids, and ortho-phthaldialdehyde (OPA) is used to react with free primary amino groups. The concentration of free amino groups can, thus, be followed using simple spectrophotometry. A mathematical model for the concentration of free primary amino groups with time is presented through which the elimination constant, and thus the half life, can be determined by curve fitting. The method is applied to inhibitors of angiotensin-converting enzyme derived from the primary structure of milk proteins. The results show that these dipeptides have in vitro half lives ranging from 4.3-64 min, when incubated with 50% rat plasma. This explains why these casokinins in vivo only cause a very moderate and short-lasting inhibition. The model for calculation of elimination constant is limited to dipeptides that do not contain a C-terminal proline. The derivatization method can be applied to longer peptides as a crude indicator of peptide hydrolysis, but does not allow calculation of their elimination constants per se.

Animals↗
PubMed2004

Bias explorer: measurements of compositional bias in EMBL and GenBank sequence files.

A Windows application for compositional analysis of sequenced genomes (EMBL or GenBank flat files) is available as freeware. The application allows the user to quantify word bias using Markov chain analysis and it allows the user to generate sliding window data for GC-skew, AT-skew, purine excess, keto excess and discrete word counts. The mathematical routines reside in a dynamic link library (DLL), which can be used independently by other applications. The software is available for download at http://www.dfuni.dk/~anfu/Bioinformatics/Main.htm.

Bias↗
PubMed2004

Bioinformatic analysis of the link between gene composition and expressivity in Saccharomyces cerevisiae and Schizosaccharomyces pombe.

The compositional non-randomness was studied in genes of Saccharomyces cerevisiae and Schizosaccharomyces pombe. In both species, codon usage is well correlated with expressivity (measured as the codon adaptation index). Both species generally display higher nucleotide non-randomness in the group of highly expressed genes than in the lowly expressed genes. The highly expressed genes in both species are furthermore characterized by marked peaks in non-randomness at N=3 upstream of start codons, N=2 downstream of start codons and at N=1 and N=7 downstream of stop codons, indicating that these nucleotides may be key elements in translational regulation. Intragenic variation in codon usage was also observed to be linked to expressivity. It is suggested that the firm link between expressivity and codon usage calls for codon optimization. Based on bioinformatic calculations, examples of proteins are given for which codon optimizations might be relevant.

Base Sequence↗
PubMed2004

Nucleotides downstream of start codons show marked non-randomness in Escherichia coli but not in Bacillus subtilis.

This study aimed at measuring the nucleotide non-randomness in the region downstream of start codons in bacterial genes and to see if the non-randomness differs between biased and unbiased genes, in terms of the effective number of codons (Nc) and the codon adaptation index (CAI). In Escherichia coli, there was a marked elevation in nucleotide conservation for the genes having low Nc-values compared to the genes having high Nc-values, i.e the more biased genes showed a higher level of non-randomness. Likewise, the genes displaying high CAI-values showed stronger nucleotide conservation than the genes of low CAI-values. This elevated conservation is visible up to approximately 15-17 nucleotides downstream of the start codon, after which there is little difference. This indicates that there may be distinct selectional mechanisms acting upon the first 5-6 codons within genes in E. coli. In B. subtilis, these effects are less pronounced, if present at all. Furthermore, analyses of codons used in this region were not in support of the hypothesis that the elevation in nucleotide non-randomness is a question of selection for certain optimal codons.

Bacillus subtilis↗
PubMed2003

Lactic acid bacteria as prime candidates for codon optimization.

In species having a strong correlation of expressivity and codon bias it has been shown that heterologous expression can be optimized by changing codons of the introduced gene towards the set of codons that the host organism naturally uses in its highly expressed genes. Even though two lactic acid bacteria are fully sequenced, there are no reports on attempts of codon optimization in the literature. In this report it is demonstrated that codons used in highly expressed genes tend to differ from the codons in lowly expressed genes, and that there is a strong correlation of codon bias and empirical expressivity (codon adaptation index) in Lactococcus lactis and Lactobacillus plantarum. This strongly suggests that codon optimization strategies could be applied to expression systems with lactic acid bacteria as producer strains. A good example of a candidate for codon optimization is the mouse interleukin-2 gene, which in its natural form has an extremely low codon adaptation index for expression in Lc. lactis.

Algorithms↗
PubMed2003

The effective number of codons for individual amino acids: some codons are more optimal than others.

The aim of this study was to evaluate the codon bias using the effective number of codons for individual amino acids (N(c)(AA)) and to assess the codon bias in relation to the definition of optimal codons, using Escherichia coli as a model organism. We show that a general correlation exists between the effective number of codons (Ncirc(c)) or codon adaptation index (CAI) and N(c)(AA), but that this correlation is not equally strong for all amino acids within a degeneracy group. For example, leucine codons contribute more to Ncirc(c) and the codon adaptation index than serine codons. A possible explanation is that some optimal codons are more optimal than others, in terms of the selectional advantage they offer. This hypothesis is confirmed by further analysis on the correlations that exist between values for relative synonymous codon usage (RSCU), N(c)(AA), and the codon adaptation index.

Amino Acids↗