The dasABC gene cluster, adjacent to dasR, encodes a novel ABC transporter for the uptake of N,N'-diacetylchitobiose in Streptomyces coelicolor A3(2).
N,N'-Diacetylchitobiose [(GlcNAc)(2)] induces the transcription of chitinase (chi) genes in Streptomyces coelicolor A3(2). Physiological studies showed that (GlcNAc)(2) addition triggered chi expression and increased the rate of (GlcNAc)(2) concentration decline in culture supernatants of mycelia already cultivated with (GlcNAc)(2), suggesting that (GlcNAc)(2) induced the synthesis of its own uptake system. Four open reading frames (SCO0531, SCO0914, SCO2946, and SCO5232) encoding putative sugar-binding proteins of ABC transporters were found in the genome by probing the 12-bp repeat sequence required for regulation of chi transcription. SCO5232, named dasA, showed transcriptional induction by (GlcNAc)(2) and N,N',N'''-triacetylchitotriose [(GlcNAc)(3)]. Surface plasmon resonance analysis showed that recombinant DasA protein exhibited the highest affinity for (GlcNAc)(2) (equilibrium dissociation constant [K(D)] = 3.22 x 10(-8)). In the dasA-null mutant, the rate of decline of the (GlcNAc)(2) concentration in the culture supernatant was about 25% of that in strain M145. The in vitro and in vivo data clearly demonstrated that dasA is involved in (GlcNAc)(2) uptake. Upstream and downstream of dasA, the transcriptional regulator gene (dasR) and two putative integral membrane protein genes (dasBC) are located in the opposite and same orientations, respectively. The expression of dasR and dasB, which seemed independent of dasA transcription, was also induced by (GlcNAc)(2) and (GlcNAc)(3).
Saito
Akihiro
A
Department of Bioresources Chemistry, Faculty of Horticulture, Chiba University, Matsudo 648, Matsudo City, Chiba 271-8510, Japan. takosaito@faculty.chiba-u.jp
Shinya
Tomonori
T
Miyamoto
Katsushiro
K
Yokoyama
Tomofumi
T
Kaku
Hanae
H
Minami
Eiichi
E
Shibuya
Naoto
N
Tsujibo
Hiroshi
H
Nagata
Yoshiho
Y
Ando
Akikazu
A
Fujii
Takeshi
T
Miyashita
Kiyotaka
K
D018528:ATP-Binding Cassette Transporters / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002688:Chitinases / Q000378:metabolism; D003470:Culture Media; D004187:Disaccharides / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D015151:Immunoblotting; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics; D016366:Open Reading Frames / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D048372:Streptomyces coelicolor / Q000235:genetics*; D020349:Surface Plasmon Resonance
D018528:ATP-Binding Cassette Transporters; D003470:Culture Media; D004187:Disaccharides; C010913:N,N-diacetylchitobiose; D002688:Chitinases