Fine structure mapping of the tryptophan genes in Pseudomonas putida.
نویسندگان
چکیده
ave used the tryptophan pathway to develop a gene transfer system via wta:sducing bacteriophage in Pseudomonas putida ( CHAKRAEARTY, GUNSALUS and GUNSALUS 1967). The tryptophan pathway itself has proved interesting in the number and nature of its enzymes, their mode of regulation, and the organization of the genes in the bacterial chromosome. Enzymatic analysis of extracts of prototrophic and auxotrophic strains grown on limiting and excess tryptophan levels has demonstrated six enzymatic activities under three types of regulation (CRAWFORD and GUNSALUS 1966). The reactions and gene designations are shown in Figure 1. Assignation of letters for the individual genes follows the suggestion of DEMEREC, et al., (1966). Three early enzymes, anthranilate synthase (AS) , phosphoribosyl transferase (PRT) , and the condensing enzyme for indole ring closure, indole-glycerol phosphate synthase (InGPS), are under repression control; they are produced in increased amounts on exhaustion of tryptophan, though not necessarily in a coordinate manner. The AS is feedback inhibited by tryptophan (QUEENER and GUNSALUS 1968). The genes controlling the synthesis of these enzymes (trpA (AS), trpB (PRT), and trpD (InGPS), are closely linked by transduction (CHAKRABARTY, GUNSALUS and GUNSALUS 1967, 1968). A first estimate of the size of the catalytically active proteins was obtained by molecular sieve filtration on Sephadex G-1 00 ( ENATSU and CRAWFORD 1968). The PRT and InGPS activities were recovered in better than 80% yield with retention volumes indicating respective molecular weights of 64,000 and 32,000. The recovery of AS activity rarely exceeds 20% and has subsequently been shown to require two protein fractions, separable from the other tryptophan enzymatic activities, with molecular sizes of about 17,000 and 80,000 estimated from a Sephadex G-100 column (QUEENER and GUNSALUS 1968). Of ten AS mutants examined, all lacked activity for the larger component. There is evidence for an intermediate between chorismate and anthranilate, based in part on the activity of the two enzyme components when separated by a dialysis membrane. SOMERVILLE and ELFORD (1967) have also presented evidence for several
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عنوان ژورنال:
- Genetics
دوره 60 3 شماره
صفحات -
تاریخ انتشار 1968