Deep learning-based prediction of protein-ligand complexes has advanced significantly with the development of architectures such as AlphaFold3, Boltz-1, Chai-1, Protenix, and NeuralPlexer. Multiple sequence alignment (MSA) has been a key input, providing coevolutionary information critical for structural inference. However, recent benchmarks reveal a major limitation: these models often memorize ligand poses from training data and perform poorly on novel chemotypes or dynamic binding events involving substantial conformational changes in binding pockets. To overcome this, we introduced a state-aware protein-ligand prediction strategy leveraging purified sequence subsets generated by AF-ClaSeq - a method previously developed by our group. AF-ClaSeq isolates coevolutionary signals and selects sequences that preferentially encode distinct structural states as predicted by AlphaFold2. By applying MSA-derived conformational restraints, we observed significant improvements in predicting ligand poses. In cases where AlphaFold3 previously failed-producing incorrect ligand placements and associated protein conformations-we were able to correct the predictions by using sequence subsets corresponding to the relevant functional state, such as the inactive form of an enzyme bound to a negative allosteric modulator. We believe this approach represents a powerful and generalizable strategy for improving protein-ligand complex predictions, with potential applications across a broad range of molecular modeling tasks.
Temporal alignment of multiple signals through time warping is crucial in many fields, such as classification within speech recognition or robot motion learning. Almost all related works are limited to data in Euclidean space. Although an attempt was made in 2011 to adapt this concept to unit quaternions, a general extension to Riemannian manifolds remains absent. Given its importance for numerous applications in robotics and beyond, we introduce Riemannian Time Warping~(RTW). This novel approach efficiently aligns multiple signals by considering the geometric structure of the Riemannian manifold in which the data is embedded. Extensive experiments on synthetic and real-world data, including tests with an LBR iiwa robot, demonstrate that RTW consistently outperforms state-of-the-art baselines in both averaging and classification tasks.
The Composition Vector Tree (CVTree) method, developed under the leadership of Professor Hao Bailin, is an alignment-free algorithm for constructing phylogenetic trees. Although initially designed for studying prokaryotic evolution based on whole-genome, it has demonstrated broad applicability across diverse biological systems and gene sequences. In this study, we employed two methods, InterList and Hao, of CVTree to investigate the phylogeny and taxonomy of prokaryote based on the 16S rRNA sequences from All-Species Living Tree Project. We have established a comprehensive phylogenetic tree that incorporates the majority of species documented in human scientific knowledge and compared it with the taxonomy of prokaryotes. And the performance of CVTree were also compared with multiple sequence alignment-based approaches. Our results revealed that CVTree methods achieve computational speeds 1-3 orders of magnitude faster than conventional alignment methods while maintaining high consistency with established taxonomic relationships, even outperforming some multiple sequence alignment methods. These findings confirm CVTree's effectiveness and efficiency not only for whole-genome evolutionary studies but also for phylogenetic and taxonomic investigations based on genes.
After AlphaFold won the Nobel Prize, protein prediction with deep learning once again became a hot topic. We comprehensively explore advanced deep learning methods applied to protein structure prediction and design. It begins by examining recent innovations in prediction architectures, with detailed discussions on improvements such as diffusion based frameworks and novel pairwise attention modules. The text analyses key components including structure generation, evaluation metrics, multiple sequence alignment processing, and network architecture, thereby illustrating the current state of the art in computational protein modelling. Subsequent chapters focus on practical applications, presenting case studies that range from individual protein predictions to complex biomolecular interactions. Strategies for enhancing prediction accuracy and integrating deep learning techniques with experimental validation are thoroughly explored. The later sections review the industry landscape of protein design, highlighting the transformative role of artificial intelligence in biotechnology and discussing emerging market trends and future challenges. Supplementary appendices provide essential resources such as databases and open source tools, making this volume a valuable reference for researchers and students.
Time Series Alignment is a critical task in signal processing with numerous real-world applications. In practice, signals often exhibit temporal shifts and scaling, making classification on raw data prone to errors. This paper introduces a novel approach for Multiple Time Series Alignment (MTSA) leveraging Deep Learning techniques. While most existing methods primarily address Multiple Sequence Alignment (MSA) for protein and DNA sequences, there remains a significant gap in alignment methodologies for numerical time series. Additionally, conventional approaches typically focus on pairwise alignment, whereas our proposed method aligns all signals in a multiple manner (all the signals are aligned together at once). This innovation not only enhances alignment efficiency but also significantly improves computational speed. By decomposing into piece-wise linear sections, we introduce varying levels of complexity into the warping function. Additionally, our method ensures the satisfaction of three warping constraints: boundary, monotonicity, and continuity conditions. The utilization of a deep convolutional network allows us to employ a new loss function, addressing some limitations of Dynamic Time Warping (DTW). Experimental results on the UCR Archive 2018, comprising 129 time series datasets, demonstrate that employing our approach to align signals significantly enhances classification accuracy and warping average and also reduces the run time across the majority of these datasets.
Deep learning approaches achieved significant progress in predicting protein structures. These methods are often applied to protein-protein interactions (PPIs) yet require Multiple Sequence Alignment (MSA) which is unavailable for various interactions, such as antibody-antigen. Computational docking methods are capable of sampling accurate complex models, but also produce thousands of invalid configurations. The design of scoring functions for identifying accurate models is a long-standing challenge. We develop a novel attention-based Graph Neural Network (GNN), ContactNet, for classifying PPI models obtained from docking algorithms into accurate and incorrect ones. When trained on docked antigen and modeled antibody structures, ContactNet doubles the accuracy of current state-of-the-art scoring functions, achieving accurate models among its Top-10 at 43% of the test cases. When applied to unbound antibodies, its Top-10 accuracy increases to 65%. This performance is achieved without MSA and the approach is applicable to other types of interactions, such as host-pathogens or general PPIs.
Sign Language Recognition (SLR) is a fast-growing field that aims to fill the communication gaps between the hearing-impaired and people without hearing loss. Existing solutions for Persian Sign Language (PSL) are limited to word-level interpretations, underscoring the need for more advanced and comprehensive solutions. Moreover, previous work on other languages mainly focuses on manipulating the neural network architectures or hardware configurations instead of benefiting from the aggregated results of multiple models. In this paper, we introduce PenSLR, a glove-based sign language system consisting of an Inertial Measurement Unit (IMU) and five flexible sensors powered by a deep learning framework capable of predicting variable-length sequences. We achieve this in an end-to-end manner by leveraging the Connectionist Temporal Classification (CTC) loss function, eliminating the need for segmentation of input signals. To further enhance its capabilities, we propose a novel ensembling technique by leveraging a multiple sequence alignment algorithm known as Star Alignment. Furthermore, we introduce a new PSL dataset, including 16 PSL signs with more than 3000 time-series samples in total. We utilize this dataset to evaluate the performance of our system based on four word-level and sentence-level metrics. Our evaluations show that PenSLR achieves a remarkable word accuracy of 94.58% and 96.70% in subject-independent and subject-dependent setups, respectively. These achievements are attributable to our ensembling algorithm, which not only boosts the word-level performance by 0.51% and 1.32% in the respective scenarios but also yields significant enhancements of 1.46% and 4.00%, respectively, in sentence-level accuracy.
Multiple Sequence Alignment (MSA) plays a pivotal role in unveiling the evolutionary trajectories of protein families. The accuracy of protein structure predictions is often compromised for protein sequences that lack sufficient homologous information to construct high quality MSA. Although various methods have been proposed to generate virtual MSA under these conditions, they fall short in comprehensively capturing the intricate coevolutionary patterns within MSA or require guidance from external oracle models. Here we introduce MSAGPT, a novel approach to prompt protein structure predictions via MSA generative pretraining in the low MSA regime. MSAGPT employs a simple yet effective 2D evolutionary positional encoding scheme to model complex evolutionary patterns. Endowed by this, its flexible 1D MSA decoding framework facilitates zero or few shot learning. Moreover, we demonstrate that leveraging the feedback from AlphaFold2 can further enhance the model capacity via Rejective Fine tuning (RFT) and Reinforcement Learning from AF2 Feedback (RLAF). Extensive experiments confirm the efficacy of MSAGPT in generating faithful virtual MSA to enhance the structure prediction accuracy. The transfer learning capabilities also highlight its great potential for facilitating other protein tasks.
Advances in gene sequencing have enabled in silico analyses of microbial genomes and have led to the revision of concepts of microbial taxonomy and evolution. We explore deficiencies in existing multiple sequence global alignment algorithms and introduce a new indexing scheme to partition the dynamic programming algorithm hypercube scoring tensor over processors based on the dependency between partitions to be scored in parallel. The performance of algorithms is compared in the study of rpoB gene sequences of Mycoplasma species.
Protein-nucleic acid interactions play a very important role in a variety of biological activities. Accurate identification of nucleic acid-binding residues is a critical step in understanding the interaction mechanisms. Although many computationally based methods have been developed to predict nucleic acid-binding residues, challenges remain. In this study, a fast and accurate sequence-based method, called ESM-NBR, is proposed. In ESM-NBR, we first use the large protein language model ESM2 to extract discriminative biological properties feature representation from protein primary sequences; then, a multi-task deep learning model composed of stacked bidirectional long short-term memory (BiLSTM) and multi-layer perceptron (MLP) networks is employed to explore common and private information of DNA- and RNA-binding residues with ESM2 feature as input. Experimental results on benchmark data sets demonstrate that the prediction performance of ESM2 feature representation comprehensively outperforms evolutionary information-based hidden Markov model (HMM) features. Meanwhile, the ESM-NBR obtains the MCC values for DNA-binding residues prediction of 0.427 and 0.391 on two independent test sets, which are 18.61 and 10.45% higher than those of the second-best methods, respectively. Moreover, by completely discarding the time-cost multiple sequence alignment process, the prediction speed of ESM-NBR far exceeds that of existing methods (5.52s for a protein sequence of length 500, which is about 16 times faster than the second-fastest method). A user-friendly standalone package and the data of ESM-NBR are freely available for academic use at: https://github.com/wwzll123/ESM-NBR.
The aim of this study was to develop a method that would identify the cluster centroids and the optimal number of clusters for a given sensitivity level and could work equally well for the different sequence datasets. A novel method that combines the linear mapping hash function and multiple sequence alignment (MSA) was developed. This method takes advantage of the already sorted by similarity sequences from the MSA output, and identifies the optimal number of clusters, clusters cut-offs, and clusters centroids that can represent reference gene vouchers for the different species. The linear mapping hash function can map an already ordered by similarity distance matrix to indices to reveal gaps in the values around which the optimal cut-offs of the different clusters can be identified. The method was evaluated using sets of closely related (16S rRNA gene sequences of Nocardia species) and highly variable (VP1 genomic region of Enterovirus 71) sequences and outperformed existing unsupervised machine learning clustering methods and dimensionality reduction methods. This method does not require prior knowledge of the number of clusters or the distance between clusters, handles clusters of different sizes and shapes, and scales linearly with the dataset. The combination of MSA with the linear mapping hash function is a computationally efficient way of gene sequence clustering and can be a valuable tool for the assessment of similarity, clustering of different microbial genomes, identifying reference sequences, and for the study of evolution of bacteria and viruses.
The translation of comparative genomics into clinical decision support tools often depends on the quality of sequence alignments. However, currently used methods of multiple sequence alignments suffer from significant biases and problems with aligning diverged sequences. The objective of this study was to develop and test a new multiple sequence alignment (MSA) algorithm suitable for the high-throughput comparative analysis of different microbial genomes. This algorithm employs an innovative tensor indexing method for partitioning the dynamic programming hyper-cube space for parallel processing. We have used the clinically relevant task of identifying regions that determine resistance to antibiotics to test the new algorithm and to compare its performance with existing MSA methods. The new method "mmDst" performed better than existing MSA algorithms for more divergent sequences because it employs a simultaneous alignment scoring recurrence, which effectively approximated the score for edge missing cell scores that fall outside the scoring region.
Generative protein language models are a natural way to design new proteins with desired functions. However, current models are either difficult to direct to produce a protein from a specific family of interest, or must be trained on a large multiple sequence alignment (MSA) from the specific family of interest, making them unable to benefit from transfer learning across families. To address this, we propose $\textbf{P}$r$\textbf{o}$tein $\textbf{E}$volutionary $\textbf{T}$ransformer (PoET), an autoregressive generative model of whole protein families that learns to generate sets of related proteins as sequences-of-sequences across tens of millions of natural protein sequence clusters. PoET can be used as a retrieval-augmented language model to generate and score arbitrary modifications conditioned on any protein family of interest, and can extrapolate from short context lengths to generalize well even for small families. This is enabled by a unique Transformer layer; we model tokens sequentially within sequences while attending between sequences order invariantly, allowing PoET to scale to context lengths beyond those used during training. In extensive experiments on deep mutational scanning datasets, we show that PoET outperforms existing protein language models and evolutionary sequence models for variant function prediction across proteins of all MSA depths. We also demonstrate PoET's ability to controllably generate new protein sequences.
This paper presents a novel evaluation approach to text-based speaker diarization (SD), tackling the limitations of traditional metrics that do not account for any contextual information in text. Two new metrics are proposed, Text-based Diarization Error Rate and Diarization F1, which perform utterance- and word-level evaluations by aligning tokens in reference and hypothesis transcripts. Our metrics encompass more types of errors compared to existing ones, allowing us to make a more comprehensive analysis in SD. To align tokens, a multiple sequence alignment algorithm is introduced that supports multiple sequences in the reference while handling high-dimensional alignment to the hypothesis using dynamic programming. Our work is packaged into two tools, align4d providing an API for our alignment algorithm and TranscribeView for visualizing and evaluating SD errors, which can greatly aid in the creation of high-quality data, fostering the advancement of dialogue systems.
With the exponential increase of the protein sequence databases over time, multiple-sequence alignment (MSA) methods, like PSI-BLAST, perform exhaustive and time-consuming database search to retrieve evolutionary information. The resulting position-specific scoring matrices (PSSMs) of such search engines represent a crucial input to many machine learning (ML) models in the field of bioinformatics and computational biology. A protein sequence is a collection of contiguous tokens or characters called amino acids (AAs). The analogy to natural language allowed us to exploit the recent advancements in the field of Natural Language Processing (NLP) and therefore transfer NLP state-of-the-art algorithms to bioinformatics. This research presents an Embedding Language Model (ELMo), converting a protein sequence to a numerical vector representation. While the original ELMo trained a 2-layer bidirectional Long Short-Term Memory (LSTMs) network following a two-path architecture, one for the forward and the second for the backward pass, by merging the idea of PSSMs with the concept of transfer-learning, this work introduces a novel bidirectional language model (bi-LM) with four times less free parameters and using rather a single path for both passes. The model was trained not only on predicting the next AA but also on the probability distribution of the next AA derived from similar, yet different sequences as summarized in a PSSM, simultaneously for multi-task learning, hence learning evolutionary information of protein sequences as well. The network architecture and the pre-trained model are made available as open source under the permissive MIT license on GitHub at https://github.com/issararab/PEvoLM.
The field of protein folding research has been greatly advanced by deep learning methods, with AlphaFold2 (AF2) demonstrating exceptional performance and atomic-level precision. As co-evolution is integral to protein structure prediction, AF2's accuracy is significantly influenced by the depth of multiple sequence alignment (MSA), which requires extensive exploration of a large protein database for similar sequences. However, not all protein sequences possess abundant homologous families, and consequently, AF2's performance can degrade on such queries, at times failing to produce meaningful results. To address this, we introduce a novel generative language model, MSA-Augmenter, which leverages protein-specific attention mechanisms and large-scale MSAs to generate useful, novel protein sequences not currently found in databases. These sequences supplement shallow MSAs, enhancing the accuracy of structural property predictions. Our experiments on CASP14 demonstrate that MSA-Augmenter can generate de novo sequences that retain co-evolutionary information from inferior MSAs, thereby improving protein structure prediction quality on top of strong AF2.
The alignment of biological sequences such as DNA, RNA, and proteins, is one of the basic tools that allow to detect evolutionary patterns, as well as functional/structural characterizations between homologous sequences in different organisms. Typically, state-of-the-art bioinformatics tools are based on profile models that assume the statistical independence of the different sites of the sequences. Over the last years, it has become increasingly clear that homologous sequences show complex patterns of long-range correlations over the primary sequence as a consequence of the natural evolution process that selects genetic variants under the constraint of preserving the functional/structural determinants of the sequence. Here, we present a new alignment algorithm based on message passing techniques that overcomes the limitations of profile models. Our method is based on a new perturbative small-coupling expansion of the free energy of the model that assumes a linear chain approximation as the $0^\mathrm{th}$-order of the expansion. We test the potentiality of the algorithm against standard competing strategies on several biological sequences.
Ancestral sequence reconstruction is a key task in computational biology. It consists in inferring a molecular sequence at an ancestral species of a known phylogeny, given descendant sequences at the tip of the tree. In addition to its many biological applications, it has played a key role in elucidating the statistical performance of phylogeny estimation methods. Here we establish a formal connection to another important bioinformatics problem, multiple sequence alignment, where one attempts to best align a collection of molecular sequences under some mismatch penalty score by inserting gaps. Our result is counter-intuitive: we show that perfect pairwise sequence alignment with high probability is possible in principle at arbitrary large evolutionary distances - provided the phylogeny is known and dense enough. We use techniques from ancestral sequence reconstruction in the taxon-rich setting together with the probabilistic analysis of sequence evolution models involving insertions and deletions.
Profile hidden Markov models (pHMMs) are widely employed in various bioinformatics applications to identify similarities between biological sequences, such as DNA or protein sequences. In pHMMs, sequences are represented as graph structures. These probabilities are subsequently used to compute the similarity score between a sequence and a pHMM graph. The Baum-Welch algorithm, a prevalent and highly accurate method, utilizes these probabilities to optimize and compute similarity scores. However, the Baum-Welch algorithm is computationally intensive, and existing solutions offer either software-only or hardware-only approaches with fixed pHMM designs. We identify an urgent need for a flexible, high-performance, and energy-efficient HW/SW co-design to address the major inefficiencies in the Baum-Welch algorithm for pHMMs. We introduce ApHMM, the first flexible acceleration framework designed to significantly reduce both computational and energy overheads associated with the Baum-Welch algorithm for pHMMs. ApHMM tackles the major inefficiencies in the Baum-Welch algorithm by 1) designing flexible hardware to accommodate various pHMM designs, 2) exploiting predictable data dependency patterns through on-chip memory with memoization techniques, 3) rapidly filtering out negligible computations using a hardware-based filter, and 4) minimizing redundant computations. ApHMM achieves substantial speedups of 15.55x - 260.03x, 1.83x - 5.34x, and 27.97x when compared to CPU, GPU, and FPGA implementations of the Baum-Welch algorithm, respectively. ApHMM outperforms state-of-the-art CPU implementations in three key bioinformatics applications: 1) error correction, 2) protein family search, and 3) multiple sequence alignment, by 1.29x - 59.94x, 1.03x - 1.75x, and 1.03x - 1.95x, respectively, while improving their energy efficiency by 64.24x - 115.46x, 1.75x, 1.96x.
Large-scale Protein Language Models (PLMs) have improved performance in protein prediction tasks, ranging from 3D structure prediction to various function predictions. In particular, AlphaFold, a ground-breaking AI system, could potentially reshape structural biology. However, the utility of the PLM module in AlphaFold, Evoformer, has not been explored beyond structure prediction. In this paper, we investigate the representation ability of three popular PLMs: ESM-1b (single sequence), MSA-Transformer (multiple sequence alignment) and Evoformer (structural), with a special focus on Evoformer. Specifically, we aim to answer the following key questions: (i) Does the Evoformer trained as part of AlphaFold produce representations amenable to predicting protein function? (ii) If yes, can Evoformer replace ESM-1b and MSA-Transformer? (ii) How much do these PLMs rely on evolution-related protein data? In this regard, are they complementary to each other? We compare these models by empirical study along with new insights and conclusions. All code and datasets for reproducibility are available at https://github.com/elttaes/Revisiting-PLMs.