Background: With more countries in the world entering elderly society, osteoporosis is a
common disease among the elderly, especially middle-aged and elderly women. Although calcitonin
is an effective drug used to treat osteoporosis in clinical practice, it also exists such problems
as high cost, short half-life, and high immunogenicity. Therefore, to explore more efficient calcitonin
has important clinical significance.
Objective: Given the emergence of new-generation gene sequencing, numerous genome sequences
of marine species have been revealed. This study aimed to identify new, highly active Calcitonins
(CTs) from the gene database.
Methods: Candidate CT sequences were obtained by BLAST and analyzed. The evolutionary tree
of these sequences was constructed using the Neighbor-Joining method of MEGA 7 software. Secondary
structures were analyzed by Circular Dichroism (CD). The biological activities of CTs were
estimated using the standard of the rat hypocalcemic activity assay in vivo. The half-life and immunogenicity
of CT sequences were determined by ELISA. The physicochemical properties of peptides
were analyzed with ProtParam and HeliQuest.
Results: A total of 64 candidate CT gene and amino acid sequences from different species were
obtained by BLAST using the salmon CT (sCT) sequence as the query sequence. These sequences
were clustered to 27 different CT polypeptide sequences, and then the evolutionary tree was constructed.
A total of 13 sequences were selected for chemical synthesis and activity assay. Results
showed that although their secondary structures were similar, four types of candidate CTs had 30%
higher activities than sCT, three other types had similar activities to sCT, and the remaining four
types had much lower activities than sCT. Among the three designed CTs, the activities of CT-01
and CT-02 were at least 50% higher than those of sCT. Furthermore, all three CT sequences had a
similar half-life to sCT and lower immunogenicity.
Conclusion: CTs from Monodelphis domestica, Gallus gallus, Ornithorhynchus anatinus, and
Carassius auratus had high activities. The exploration and mining of the marine-life genome database
can be extremely valuable considering broad application prospect.