The amino acid sequences of proteins determine their three-dimensional structures and functions. However, how sequence information is related to structures and functions is still enigmatic. In this study, we show that at least a part of the sequence information can be extracted by treating amino acid sequences of proteins as a collection of English words, based on a working hypothesis that amino acid sequences of proteins are composed of short constituent amino acid sequences (SCSs) or “words”. We first confirmed that the English language highly likely follows Zipf's law, a special case of power law. We found that the rank-frequency plot of SCSs in proteins exhibits a similar distribution when low-rank tails are excluded. In comparison with natural English and “compressed” English without spaces between words, amino acid sequences of proteins show larger linear ranges and smaller exponents with heavier low-rank tails, demonstrating that the SCS distribution in proteins is largely scale-free. A distribution pattern of SCSs in proteins is similar among species, but species-specific features are also present. Based on the availability scores of SCSs, we found that sequence motifs are enriched in high-availability sites (i.e., “key words”) and vice versa. In fact, the highest availability peak within a given protein sequence often directly corresponds to a sequence motif. The amino acid composition of high-availability sites within motifs is different from that of entire motifs and all protein sequences, suggesting the possible functional importance of specific SCSs and their compositional amino acids within motifs. We anticipate that our availability-based word decoding approach is complementary to sequence alignment approaches in predicting functionally important sites of unknown proteins from their amino acid sequences.

Protein structure and function information is coded in amino acid sequences. However, the relationship between primary sequences and three-dimensional structures and functions remains enigmatic. Our approach to this fundamental biochemistry problem is based on the frequencies of short constituent sequences (SCSs) or words. A protein amino acid sequence is considered analogous to an English sentence, where SCSs are equivalent to words. Availability scores, which are defined as real SCS frequencies in the non-redundant amino acid database relative to their probabilistically expected frequencies, demonstrate the biological usage bias of SCSs. As a result, this frequency-based linguistic approach is expected to have diverse applications, such as secondary structure specifications by structure-specific SCSs and immunological adjuvants with rare or non-existent SCSs. Linguistic similarities (e.g., wide ranges of scale-free distributions) and dissimilarities (e.g., behaviors of low-rank samples) between proteins and the natural English language have been revealed in the rank-frequency relationships of SCSs or words. We have developed a web server, the SCS Package, which contains five applications for analyzing protein sequences based on the linguistic concept. These tools have the potential to assist researchers in deciphering structurally and functionally important protein sites, species-specific sequences, and functional relationships between SCSs. The SCS Package also provides researchers with a tool to construct amino acid sequences de novo based on the idiomatic usage of SCSs.

Little is known about protein sequences unique in humans. Here, we performed alignment-free sequence comparisons based on the availability (frequency bias) of short constituent amino acid (aa) sequences (SCSs) in proteins to search for human-specific proteins. Focusing on 5-aa SCSs (pentats), exhaustive comparisons of availability scores among the human proteome and other nine mammalian proteomes in the nonredundant (nr) database identified a candidate protein containing WRWSH, here called FAM75, as human-specific. Examination of various human genome sequences revealed that FAM75 had genomic DNA sequences for either WRWSH or WRWSR due to a single nucleotide polymorphism (SNP). FAM75 and its related protein FAM205A were found to be produced through alternative splicing. The FAM75 transcript was found only in humans, but the FAM205A transcript was also present in other mammals. In humans, both FAM75 and FAM205A were expressed specifically in testis at the mRNA level, and they were immunohistochemically located in cells in seminiferous ducts and in acrosomes in spermatids at the protein level, suggesting their possible function in sperm development and fertilization. This study highlights a practical application of SCSbased methods for protein searches and suggests possible contributions of SNP variants and alternative splicing of FAM75 to human evolution.

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