Mitomycins biosynthesis

Neither shikimic acid66 nor its methyl ester, nor dehydroquinic acid,65 were precursors of mitomycin, so it is likely that deviation to mitomycin biosynthesis occurs from DAHP (150) via, it has been suggested,65 (151), as shown in Scheme 12. 

Molecular genetic manipulation of the mitomycin pathway can elucidate the sequence of reactions involved in mitomycin biosynthesis, as well as provide access to novel mitomycin natural products. Thus, 9a-demethyl mitomycin A (32), 9-epi-mitomycin B (33), and A -methylmitomycin A (mitomycin F, 29) have been obtained using mitomycin B as starting material [69]. Mitomycin J (35) and mitomycin D (36) were isolated as minor antibiotics fromStreptomycesfradiae SCF5 [70], and mitomycin E (37) was obtained from S. lavendulae [71]. Mitomycin C, A, and F showed... 

Fig. 4. Kinetics of ethionine inhibition of mitomycin biosynthesis in synthetic medium. Numbered arrows refer to the introduction of lOOmcg/ml of D,L-ethionine to the fermentation. Corresponding numbers in parenthesis designate the plot of antibiotic synthesis resulting from such additions. Curve 6 is the plot of uninhibited antibiotic...

Fig. 5- Reversal of ethionine-inhibited mitomycin biosynthesis by L-methionine...

Fig. 7. Relationship of time of D,L-ethionine and L-methionine addition on reversal of inhibited mitomycin biosynthesis...

A number of mitomycin analogues have been prepared by precursor-directed biosynthesis [104]. A range of amines were fed to S. caespitosus, and novel derivatives of mitomycin C (type I analogues) and mitomycin B (type II analogues) were identified and in some cases (42-46 and 52-56 Scheme 11.4) isolated and characterized. Antibiotic and antitumor activities were comparable to those of mitomycin C, with the type I analogues more active than the type II analogues. 

Figure 11.9 Role of the MitM methyltransferase in mitomycin C biosynthesis.

There has been one report on the biosynthesis of FR900482 [114]. Radiolabeled D-glucosamine 33 and AHBA 38 were efficiently incorporated into 6, and D-[l-13C]-glucosamine was incorporated in the expected orientation (Figure 11.10). The biosynthetic gene cluster for FR900482 has been isolated from Streptomyces sandaensis and indicates a strong biosynthetic relationship with mitomycin C (Y. Mao, D. H. Sherman, unpublished results). 

The antibiotic mitomycin C, introduced to give a concentration of 10 jLtg. per ml. in the perfusing blood at the outset, and a continuous infusion of a total of 1 mg. over the course of 4 to 5 hours, was also associated with significant suppression of fibrinogen biosynthesis. The concentrations of mitomycin C used here are at least ten times that which significantly inhibited cellular multiplication in isolated cell cultures. Further studies at lower concentration levels with both puromy-cin and mitomycin C are indicated. 

The view that this is true net biosynthesis of fibrinogen is supported by a variety of ancillary observations. Fibrinogen biosynthesis is suppressed in the presence of metabolic analogs, such as L-ethionine and puromycin, most markedly by the latter in spite of the maximal stimulus for production. Mitomycin C, which is believed to interfere with biosynthetic processes in the nucleus, also caused some suppression of fibrinogen biosynthesis. The isolated perfused liver in the presence of any of the three inhibitors used continues to function in an apparently normal manner in terms of bile secretion, linear urea production, amino acid oxidation, and glucose utilization. The effects of these inhibitors on the biosynthesis of the other plasma proteins will be described elsewhere. 

The evidence then is that, for rifamycin and other ansamycins, biosynthesis diverts at a so-far unidentified (but early) compound in the shikimic acid pathway to give 3-amino-5-hydroxybenzoic acid (91) (as its CoA ester). This compound then yields, on the one hand, the mitomycins [e.g. porfiromycin (88)1 and, on the other, the CoA ester of P8/1-OG (92), which then affords diverse metabolites such as rifamycin B (87) and actamycin (86) (cf. ref. 83 for a detailed scheme). 

One classical and early example of selective inhibition of DNA biosynthesis is shown in Fig. 3 for the anitbiotic, mitomycin C29 A concentration of 0.1 pg/ml (3 x 10 7 M) completely inhibited DNA synthesis in E. coli B, while RNA synthesis, protein synthesis and growth meaning turbidity, i.e. cell mass increase, continued. However, after the experimental period of only 90 min, the number of viable bacteria had decreased by 85 per cent. By that time, bacterial filaments were visible under the microscope. 

Streptovaridns and Mitomycins.—Consideration of the structures deduced for metabolites with the streptovaricin skeleton which lack in particular the extensive oxygenation of say streptovaricin D, has allowed a reasonable sequence of events for streptovaricin biosynthesis to be proposed. (The streptovaridns belong to the class of ansamydns. For earlier reviews on ansamycin biosynthesis see ref. 5, p. 52, and ref. 6, p. 45.)... 

Intensive study of mitomycin (18) biosynthesis 56 has not resolved all the problems of its origin. D-Glucosamine provides carbon and nitrogen for the aziridine ring and possibly a CgN unit joins with a C7N unit of less certain origin in the biosynthesis. The latter may be related to some intermediate of the shikimate pathway 57 and intermediates of the arginine biosynthetic pathway contribute the carbamoyl group 59,57, 60, 58t... 

Geldanomycin, Rif.amycin, and Antibiotic A23187. — It is now clear that 3-amino-5-hydroxybenzoic acid (103) is the long-sought, key CyN intermediate involved in the biosynthesis of rifamycins and mitomycins. On the other hand, the C N intermediate involved in the biosynthesis of the 3-aminoacetophenone residue in pactamycin (101) is 3-aminobenzoic acid (100) (cf. Vol. 12, p.21). 

Mitomydn and Rifamycin.—Common to antibiotics like mitomycin C (190) and rifamycin S (193) is a C7N unit (shown in heavy bonding) which, it has been supposed, derives from carbohydrate metabolism. Recent experiments on mitomycin and rifamycin biosynthesis have been directed to discovering more precisely what the intermediates in the generation of this unit are. 

The w-CyN unit described was shown to be incorporated in the biosynthesis of both the ansamycins and also other related antibiotics, such as pactamycin and other compounds [188-190]. With respect to other naturally occurring compounds which contain a partial structure derived from the m-CyN unit in the molecule, asukamycin [191] is a possible shunt metabolite from 3-dehydroquinic acid in the shikimate pathway. Incorporation of [l - 3C]-AHBA (118) into the C-6 methyl group of porfiromycin (120) (a mitomycin group antibiotic) was reported (Fig. 6) [192], and the participation of a m-CyN unit in this biosynthetic pathway and the antibiotics containing a m-CyN unit were reviewed [178,188,190]. 

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