• tobacco calmodulin;
  • infrared spectroscopy;
  • secondary structure;
  • coordination structure

Calmodulin (CaM) is a Ca2+-binding protein that regulates a number of fundamental cellular activities. Nicotiana tabacum CaM (NtCaM) comprises 13 genes classified into three types, among which gene expression and target enzyme activation differ. We performed Fourier-transform infrared spectroscopy to compare the secondary and coordination structures of Mg2+ and Ca2+ among NtCaM1, NtCaM3, and NtCaM13 as representatives of the three types of NtCaMs. Data suggested that NtCaM13 has a different secondary structure due to the weak β-strand bands and the weak 1661 cm−1 band. Coordination structures of Mg2+ of NtCaM3 and NtCaM13 were similar but different from that of NtCaM1, while the Ca2+-binding manner was similar among the three CaMs. The amplitude differences of the band at 1554–1550 cm−1 obtained by second-derivative spectra indicated that the intensity change of the band of NtCaM13 was smaller in response to [Ca2+] increases under low [Ca2+] conditions than were those of NtCaM1 and NtCaM3, while the intensity reached the same level under high [Ca2+]. Therefore, NtCaM13 has a characteristic secondary structure and specific Mg2+-binding manner and needs higher [Ca2+] for bidentate Ca2+ coordination of 12th Glu in EF-hand motifs. The Ca2+-binding mechanisms of the EF-hand motifs of the three CaMs are similar; however, the cation-dependent conformational change in NtCaM13 is unique among the three NtCaMs. © 2013 Wiley Periodicals, Inc. Biopolymers 99: 472–483, 2013.