Cloning and characterization of CalS7 from Micromonospora echinospora sp. calichensis as a glucose-1-phosphate nucleotidyltransferase.

Dinesh Simkhada, Tae-Jin Oh, Eui Min Kim, Jin Cheol Yoo, Jae Kyung Sohng

Institute of Biomolecule Reconstruction (iBR), Department of Pharmaceutical Engineering, Sun Moon University, # 100 Kalsan-ri, Tangjeong-myeon, Asansi, Chungnam 336-708, Republic of Korea.

Biotechnology letters 2009 Jan

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The deoxysugar biosynthetic gene cluster of calicheamicin contains the calS7, which encodes glucose-1-phosphate nucleotidyltransferase and converts glucose-1-phosphate and nucleotides (NTP) to NDP-glucose and pyrophosphate. calS7 was expressed in Escherichia coli BL21(DE3), and the purified protein had significant thymidylyltransferase and uridylyltransferase activities as well, with some guanidylyltransferase activity but negligible cytidyl and adenyltransferase activity. The functions of thymidylyltransferase and uridylyltransferase were also verified using one-pot enzymatic synthesis of TMK and ACK. The products were analyzed by HPLC and ESI/MS, which showed peaks at m/z = 563 and 565 for TDP-D: -glucose and UDP-D-glucose, respectively, in negative mode.

Citation

Dinesh Simkhada, Tae-Jin Oh, Eui Min Kim, Jin Cheol Yoo, Jae Kyung Sohng. Cloning and characterization of CalS7 from Micromonospora echinospora sp. calichensis as a glucose-1-phosphate nucleotidyltransferase. Biotechnology letters. 2009 Jan;31(1):147-53