Pairwise alignment incorporating dipeptide covariation

Motivation: Standard algorithms for pairwise protein sequence alignment make the simplifying assumption that amino acid substitutions at neighboring sites are uncorrelated. This assumption allows implementation of fast algorithms for pairwise sequence alignment, but it ignores information that could conceivably increase the power of remote homolog detection. We examine the validity of this assumption by constructing extended substitution matrixes that encapsulate the observed correlations between neighboring sites, by developing an efficient and rigorous algorithm for pairwise protein sequence alignment that incorporates these local substitution correlations, and by assessing the ability of this algorithm to detect remote homologies. Results: Our analysis indicates that local correlations between substitutions are not strong on the average. Furthermore, incorporating local substitution correlations into pairwise alignment did not lead to a statistically significant improvement in remote homology detection. Therefore, the standard assumption that individual residues within protein sequences evolve independently of neighboring positions appears to be an efficient and appropriate approximation

Parametric Alignment of Drosophila Genomes

The classic algorithms of Needleman--Wunsch and Smith--Waterman find a maximum a posteriori probability alignment for a pair hidden Markov model (PHMM). In order to process large genomes that have undergone complex genome rearrangements, almost all existing whole genome alignment methods apply fast heuristics to divide genomes into small pieces which are suitable for Needleman--Wunsch alignment. In these alignment methods, it is standard practice to fix the parameters and to produce a single alignment for subsequent analysis by biologists. Our main result is the construction of a whole genome parametric alignment of Drosophila melanogaster and Drosophila pseudoobscura. Parametric alignment resolves the issue of robustness to changes in parameters by finding all optimal alignments for all possible parameters in a PHMM. Our alignment draws on existing heuristics for dividing whole genomes into small pieces for alignment, and it relies on advances we have made in computing convex polytopes that allow us to parametrically align non-coding regions using biologically realistic models. We demonstrate the utility of our parametric alignment for biological inference by showing that cis-regulatory elements are more conserved between Drosophila melanogaster and Drosophila pseudoobscura than previously thought. We also show how whole genome parametric alignment can be used to quantitatively assess the dependence of branch length estimates on alignment parameters. The alignment polytopes, software, and supplementary material can be downloaded at http://bio.math.berkeley.edu/parametric/.Comment: 19 pages, 3 figure

Statistical power of phylo-HMM for evolutionarily conserved element detection

<p>Abstract</p> <p>Background</p> <p>An important goal of comparative genomics is the identification of functional elements through conservation analysis. Phylo-HMM was recently introduced to detect conserved elements based on multiple genome alignments, but the method has not been rigorously evaluated.</p> <p>Results</p> <p>We report here a simulation study to investigate the power of phylo-HMM. We show that the power of the phylo-HMM approach depends on many factors, the most important being the number of species-specific genomes used and evolutionary distances between pairs of species. This finding is consistent with results reported by other groups for simpler comparative genomics models. In addition, the conservation ratio of conserved elements and the expected length of the conserved elements are also major factors. In contrast, the influence of the topology and the nucleotide substitution model are relatively minor factors.</p> <p>Conclusion</p> <p>Our results provide for general guidelines on how to select the number of genomes and their evolutionary distance in comparative genomics studies, as well as the level of power we can expect under different parameter settings.</p

A Probabilistic Model of Local Sequence Alignment That Simplifies Statistical Significance Estimation

Sequence database searches require accurate estimation of the statistical significance of scores. Optimal local sequence alignment scores follow Gumbel distributions, but determining an important parameter of the distribution (λ) requires time-consuming computational simulation. Moreover, optimal alignment scores are less powerful than probabilistic scores that integrate over alignment uncertainty (“Forward” scores), but the expected distribution of Forward scores remains unknown. Here, I conjecture that both expected score distributions have simple, predictable forms when full probabilistic modeling methods are used. For a probabilistic model of local sequence alignment, optimal alignment bit scores (“Viterbi” scores) are Gumbel-distributed with constant λ = log 2, and the high scoring tail of Forward scores is exponential with the same constant λ. Simulation studies support these conjectures over a wide range of profile/sequence comparisons, using 9,318 profile-hidden Markov models from the Pfam database. This enables efficient and accurate determination of expectation values (E-values) for both Viterbi and Forward scores for probabilistic local alignments

Improvements in the Accuracy of Pairwise Genomic Alignment

Pairwise sequence alignment is a fundamental problem in bioinformatics with wide applicability. This thesis presents three new algorithms for this well-studied problem. First, we present a new algorithm, RDA, which aligns sequences in small segments, rather than by individual bases. Then, we present two algorithms for aligning long genomic sequences: CAPE, a pairwise global aligner, and FEAST, a pairwise local aligner. RDA produces interesting alignments that can be substantially different in structure than traditional alignments. It is also better than traditional alignment at the task of homology detection. However, its main negative is a very slow run time. Further, although it produces alignments with different structure, it is not clear if the differences have a practical value in genomic research. Our main success comes from our local aligner, FEAST. We describe two main improvements: a new more descriptive model of evolution, and a new local extension algorithm that considers all possible evolutionary histories rather than only the most likely. Our new model of evolution provides for improved alignment accuracy, and substantially improved parameter training. In particular, we produce a new parameter set for aligning human and mouse sequences that properly describes regions of weak similarity and regions of strong similarity. The second result is our new extension algorithm. Depending on heuristic settings, our new algorithm can provide for more sensitivity than existing extension algorithms, more specificity, or a combination of the two. By comparing to CAPE, our global aligner, we find that the sensitivity increase provided by our local extension algorithm is so substantial that it outperforms CAPE on sequence with 0.9 or more expected substitutions per site. CAPE itself gives improved sensitivity for sequence with 0.7 or more expected substitutions per site, but at a great run time cost. FEAST and our local extension algorithm improves on this too, the run time is only slightly slower than existing local alignment algorithms and asymptotically the same

Finite sample sizes and phylogeny do not ACCOUNT FOR THE MUTUAL INFORMATION OBSERVED FOR MOST SITE-PAIRS IN MULTIPLE SEQUENCE ALIGNMENT

Mutual information (MI) is a measure frequently used to find co-evolving sites in protein families. However, factors unrelated to protein structure and function, in particular sampling variance in amino acid counts and complex evolutionary relationships among sequences, contribute to ML Understanding the contribution of these components is essential for isolating the MI associated with structural or functional co-evolution. To date, the contributions of these factors to mutual information have not been fully elucidated. We find that stochastic variations in amino acid counts and shared phylogeny each contribute substantially to measured MI. Nonetheless, the mutual information observed in real-world protein families is consistently higher than the expected contribution of these two factors. In contrast, when using synthetic data with realistic substitution rates and phylogenies, but without structural or functional constraints, the observed levels of MI match those expected due to stochastic and phylogenetic background. Our results suggest that either low levels of co-evolution are ubiquitous across positions in protein families, or some unknown factor exists beyond the currently hypothesized components of intra-protein mutual information: sampling variance, phylogenetics and structural/functional co-evolution

Generalized affine gap costs for protein sequence alignment

ABSTRACT Based on the observation that a single mutational event can delete or insert multiple residues, affine gap costs for sequence alignment charge a penalty for the existence of a gap, and a further length-dependent penalty. From structural or multiple alignments of distantly related proteins, it has been observed that conserved residues frequently fall into ungapped blocks separated by relatively nonconserved regions. To take advantage of this structure, a simple generalization of affine gap costs is proposed that allows nonconserved regions to be effectively ignored. The distribution of scores from local alignments using these generalized gap costs is shown empirically to follow an extreme value distribution. Examples are presented for which generalized affine gap costs yield superior alignments from the standpoints both of statistical significance and of alignment accuracy. Guidelines for selecting generalized affine gap costs are discussed, as is their possible application to multiple alignment. Proteins 32:88-96, 1998. 1998 Wiley-Liss, Inc.

Computational Molecular Coevolution

A major goal in computational biochemistry is to obtain three-dimensional structure information from protein sequence. Coevolution represents a biological mechanism through which structural information can be obtained from a family of protein sequences. Evolutionary relationships within a family of protein sequences are revealed through sequence alignment. Statistical analyses of these sequence alignments reveals positions in the protein family that covary, and thus appear to be dependent on one another throughout the evolution of the protein family. These covarying positions are inferred to be coevolving via one of two biological mechanisms, both of which imply that coevolution is facilitated by inter-residue contact. Thus, high-quality multiple sequence alignments and robust coevolution-inferring statistics can produce structural information from sequence alone. This work characterizes the relationship between coevolution statistics and sequence alignments and highlights the implicit assumptions and caveats associated with coevolutionary inference. An investigation of sequence alignment quality and coevolutionary-inference methods revealed that such methods are very sensitive to the systematic misalignments discovered in public databases. However, repairing the misalignments in such alignments restores the predictive power of coevolution statistics. To overcome the sensitivity to misalignments, two novel coevolution-inferring statistics were developed that show increased contact prediction accuracy, especially in alignments that contain misalignments. These new statistics were developed into a suite of coevolution tools, the MIpToolset. Because systematic misalignments produce a distinctive pattern when analyzed by coevolution-inferring statistics, a new method for detecting systematic misalignments was created to exploit this phenomenon. This new method called ``local covariation\u27\u27 was used to analyze publicly-available multiple sequence alignment databases. Local covariation detected putative misalignments in a database designed to benchmark sequence alignment software accuracy. Local covariation was incorporated into a new software tool, LoCo, which displays regions of potential misalignment during alignment editing assists in their correction. This work represents advances in multiple sequence alignment creation and coevolutionary inference

Noncoding RNA gene detection using comparative sequence analysis

BACKGROUND: Noncoding RNA genes produce transcripts that exert their function without ever producing proteins. Noncoding RNA gene sequences do not have strong statistical signals, unlike protein coding genes. A reliable general purpose computational genefinder for noncoding RNA genes has been elusive. RESULTS: We describe a comparative sequence analysis algorithm for detecting novel structural RNA genes. The key idea is to test the pattern of substitutions observed in a pairwise alignment of two homologous sequences. A conserved coding region tends to show a pattern of synonymous substitutions, whereas a conserved structural RNA tends to show a pattern of compensatory mutations consistent with some base-paired secondary structure. We formalize this intuition using three probabilistic "pair-grammars": a pair stochastic context free grammar modeling alignments constrained by structural RNA evolution, a pair hidden Markov model modeling alignments constrained by coding sequence evolution, and a pair hidden Markov model modeling a null hypothesis of position-independent evolution. Given an input pairwise sequence alignment (e.g. from a BLASTN comparison of two related genomes) we classify the alignment into the coding, RNA, or null class according to the posterior probability of each class. CONCLUSIONS: We have implemented this approach as a program, QRNA, which we consider to be a prototype structural noncoding RNA genefinder. Tests suggest that this approach detects noncoding RNA genes with a fair degree of reliability