The pur3 gene from the pur cluster encodes a monophosphatase essential for puromycin biosynthesis in Streptomyces.
The pur3 gene of the puromycin (pur) cluster from Streptomyces alboniger is essential for the biosynthesis of this antibiotic. Cell extracts from Streptomyces lividans containing pur3 had monophosphatase activity versus a variety of mononucleotides including 3'-amino-3'-dAMP (3'-N-3'-dAMP), (N6,N6)-dimethyl-3'-amino-3'-dAMP (PAN-5'-P) and AMP. This is in accordance with the high similarity of this protein to inositol monophosphatases from different sources. Pur3 was expressed in Escherichia coli as a recombinant protein and purified to apparent homogeneity. Similar to the intact protein in S. lividans, this recombinant enzyme dephosphorylated a wide variety of substrates for which the lowest Km values were obtained for the putative intermediates of the puromycin biosynthetic pathway 3'-N-3'-dAMP (Km = 1.37 mM) and PAN-5'-P (Km = 1.40 mM). The identification of this activity has allowed the revision of a previous proposal for the puromycin biosynthetic pathway.
Sánchez
María Blanca
MB
Centro de Biología Molecular Severo Ochoa, Departamento de Biología Molecular (CSIC-UAM), Universidad Autónoma Madrid, Cantoblanco, 28049 Madrid, Spain.
Barrado
Patricia
P
Jiménez
Antonio
A
Fernández Lobato
María
M
D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D005810:Multigene Family; D009711:Nucleotides / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics*; D011691:Puromycin / Q000096:biosynthesis*; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics*; D048170:Streptomyces lividans; D013379:Substrate Specificity
D009711:Nucleotides; D011691:Puromycin; D010744:Phosphoric Monoester Hydrolases