Abstract: In order to reveal the function of carotenoid cleavage dioxygenases (CCDs) in the growth and development of tobacco plants, a CCD4 (NtCCD4) gene was cloned from Samsun oriental tobacco by RT-PCR. According to sequence analysis, with a coding sequence of 1806 bp in length the gene encoded 601 amino acids. The sequence was submitted to NCBI and its GenBank accession number was JF947192. Various bioinformatics softwares were adopted to analyze and predict physicochemical properties, advanced structure and biological function of NtCCD4 protein. The results showed that the molecular weight and isoelectric point of the encoded protein were 65936.3 Da and 6.58, respectively. The protein located on chloroplast membrane and was a stable and hydrophilic protein. The secondary structure prediction indicated that the protein structure was composed mainly of random coil (57.4%) and alpha helixes (13.31%), beta turn (6.61%). The phylogenetic tree analysis showed that NtCCD4 belonged to CCDs family containing numerous conserved domains, of which the 4 histidine residues were necessary for composing the functional domain and constructed the active center of the protein by combining with Fe²⁺.