Fingerprinting of Proteins that Mediate Quagga Mussel Adhesion using a De Novo Assembled Foot Transcriptome

Abstract

The European freshwater mollusk Dreissena bugensis (quagga mussel), an invasive species to North America, adheres to surfaces underwater via the byssus: a non-living protein 'anchor'. In spite of its importance as a biofouling species, the sequence of the majority of byssal proteins responsible for adhesion are not known, and little genomic data is available. To determine protein sequence information, we utilized next-generation RNA sequencing and de novo assembly to construct a cDNA library of the quagga mussel foot transcriptome, which contains over 200,000 transcripts. Quagga mussel byssal proteins were extracted from freshly induced secretions and analyzed using LC-MS/MS; peptide spectra were matched to the transcriptome to fingerprint the entire protein primary sequences. We present the full sequences of fourteen novel quagga mussel byssal proteins, named Dreissena bugensis foot proteins 4 to 17 (Dbfp4-Dbfp17), and new sequence data for two previously observed byssal proteins Dbfp1 and Dbfp2. Theoretical masses of the newly discovered proteins range from 4.3 kDa to 21.6 kDa. These protein sequences are unique but contain features similar to glue proteins from other species, including a high degree of polymorphism, proteins with repeated peptide motifs, disordered protein structure, and block structures.

Figure 1

Dbfp7 variants with signal peptide removed, aligned using Clustal to illustrate the effects of alternative splicing. Greek letters replaced by English letters. Stars beneath sequences indicate exact match, a colon dot indicates substitution with strongly similar properties, and a single dot indicates a substitution with weakly similar properties.

Figure 2

Dbfp2 assembly, with tandem repeats highlighted in green, yellow, and blue.

Figure 3

Kyte-Doolittle hydropathy plot (right) for Dbfp4 indicating alternating domains; higher score indicates higher hydrophobicity. Hydrophobic residues are highlighted in red (left).

Figure 4

Dbfp5 protein with interesting repeats highlighted (left), and IUPred disorder prediction chart for Dbfp5, where values above 0.5 indicate a tendency to be disordered (right). Note how the cysteine-containing region has low disorder.

Figure 5

IUPred Disorder prediction chart for Dbfp13α, where values above 0.5 indicate a tendency to be disordered.