RVDB-prot, a reference viral protein database and its HMM profiles

Conversion from RVDB nucleic database to RVDB-prot

The current version of RVDB, v19.0⁴ consists of a collection of 3 084 319 nucleic sequences². The accession numbers were extracted in order to gather the corresponding database entries in Genbank format. From these entries, the corresponding coding domain protein sequences, description, and protein accession numbers were automatically recognized and copied into the protein collection. The process relies on the amino-acid sequences and information provided initially in the nucleic entry annotations. The resulting protein file contains the nucleic sequence reference, for traceability purposes. The sequence names are formatted in the following way:

>acc|<p_bank>|<p_acc>|<n_bank>|<n_acc>|<descr[sp]>, where:

p_bank is the bank in which the protein can be found

p_acc is the accession number corresponding to the protein sequence

n_bank is the bank in which the original nucleotide sequence was found

n_acc is the original information found in the nucleic database

descr is the description of the protein sequence as found in the database entry

sp is the species name.

This process produces a 4 705 359 protein sequence file.

Generation of HMM profiles

The HMM generation rationale was inspired from vFam (the database of HMM profiles built from all the viral proteins present in RefSeq, discontinued from 2014)⁵, but was entirely re-coded as a Snakemake pipeline⁶, using different tools for some key steps (clustering, alignment). The proteins sequences were clustered with a 100% identity criterion to remove duplicates using CD-Hit 4.7.0⁷. Then, the sequences were processed using Blast 2.2.26⁸ performing an all-against-all comparison. These comparisons allowed Silix 1.2.6⁹ (using default parameters) to define clusters of sequences according to their similarity. This step produced a text file in which each sequence was associated to one cluster. The information of each cluster (containing at least four sequences) was transformed into a fasta file containing all the sequences within the cluster. Then, sequences were aligned using Mafft 7.023¹⁰ in auto mode. The multiple sequence alignments were processed by HMMER 3.2.1³ (hmmbuild, using default parameters) in order to obtain the HMM profiles. The HMM profiles were finally grouped into a single file.

Annotation of HMM profiles

A cluster is defined as a set of sequences, among which each sequence is characterized by its taxonomy (i.e. a virus species) and eis associated with a description of its putative function, when it is known. In order to describe the different clusters, these information and other indicators (such as the cluster length and number of sequences) are combined into an annotation database, in SQLite format. The schema of this database is shown in Figure 1.

Figure 1. Schema of the annotation database.

The first type of data associated to a cluster is a set of keywords describing the putative function of the proteins present in a given cluster. These keywords correspond to the union of all names of the significant sequences found in PFAM¹¹ (with --cut_ga parameter which tells HMMER to trust the cutoff defined by PFAM) using all the sequences of the cluster as queries, weighted according to their frequencies, and excluding trivial words. We also produce a complementary word frequency count using sequence descriptions. These keywords are stored separately from the PFAM ones. Despite the fact that sequence descriptions can be vague or inaccurate, they are a good fallback in case the cluster had no match with any PFAM one.

Here is an example of different keywords, using annotations from the cluster number 1, containing 18 sequences; the keywords and their frequencies from PFAM, are: RNA(58), viral(18), dependent(18), polymerase(18), helicase(17), cysteine(11), picorna(11); the first ten keywords from the sequence descriptions are: virus(17), protein(13), hypothetical(10), like(9), picorna(9), polyprotein(5), RNA(4), Wenzhou(4), Beihai(4), non-structural(2). Altogether, these keywords allow to describe a cluster composed of RNA-dependent RNA polymerase of picorna-like viruses. The complementarity of these two annotations is well illustrated here since the simple list of keywords would not have allowed to identify the function of this cluster (here the viral polymerase) without PFAM.

In addition to the protein description, the database stores the virus taxonomy associated to all the taxa, referring to tNCBI TaxIDs. For each cluster, the taxonomic information is summarized by a Last Common Ancestor (LCA) that corresponds to the taxon in the tree of life to which all the sequence taxa belong; this LCA can be close to the root of the tree (Viruses), but is usually specific to a family.

Finally, the database also provides the length (number of amino acids of the multiple sequences alignment) and the number of sequences in each cluster.

This database is available in SQLite format, and to provide more direct access, flat text files are proposed. A text file for each cluster, identified with its cluster number, contains all the information related to it.

Underlying data

Database files are available at https://rvdb-prot.pasteur.fr/. Release 19.0 described in this manuscript is also available from Zenodo.

Zenodo: U-RVDBv19.0 https://doi.org/10.5281/zenodo.4002051⁴.

This project contains the following underlying data:

Data are available under the terms of the Creative Commons Attribution 4.0 International license (CC-BY 4.0).

Table 1 shows some summary metrics for the entries of this release and the different resources.

Updates are manually curated each time a new release of the main database (nucleic RVDB) is announced, i.e., four times a year. The following older versions are also available online: 18.0 (2020–03), 17.0 (2019–11), 16.0 (2019–06), 15.1 (2019–02), 14.0 (2018–09), 13.0 (2018–06), 12.2(2018–03), 11.5 (2017-–0), 10.2 (2017–04).

Usage HMMER can be used to search for all profiles in a fasta sequence file (sequences.fasta): hmmsearch U-RVDBv15.1-prot.fasta-prot.hmm sequences.fasta > result.out. Additional options are available in HMMER User’s Guide.