Kinetics, alkaline elution

Figure 3, The alkaline elution kinetics of DNA from L1210 cells treated with cis-DDP (10 jM.) for 1 h followed by 12 h incubation in a drug-free medium, L1210 cells were labeled with C-thymidine for 20 h and were either cis-DDP treated (A A) or untreated (%, O). (Reproduced with permission from Ref. 20, Copyright 1980, Academic Press.)...

The scheme in Figure 1 has implicit within it a kinetic approach to the study of DNA damage and repair and its relationship to desirable and toxic consequences of antineo-plastic, DNA reactive drugs In this paper the stepwise approach to the study of this scheme for one agent, cis-DDP, is presented. The initial selection of cis-DDP was made because of its clinical efficacy and the availability of an Inactive isomer Q). We had hoped that some critical DNA lesion would be more efficiently produced by the active compound The development of the alkaline elution technique incorporating enzymatic deproteinization to distinguish between interstrand and DNA-protein crosslinking was critical for this analysis ) ... 

Lakshmi and Balasubramanian (1980) showed the presence of a new multiple form of arylsulfohydrolase B in human and monkey brain. Arylsulfohydrolase B, can be separated by DEAE-cellulose chromatography (Mathew and Balasubramanian, 1984). The B, form totally binds to Sephadex G-200 and was not eluted with 1.0 M NaCl, 0.5 M glucose, 0.5 M glucose plus 0.5 M NaCl, 0.5 M KSCN, 1 M urea, or 1% Triton X-100. The treatment of arylsulfohydrolase B with Escherichia coli alkaline phosphatase results in the formation of a less acidic form, presumably due to dephosphorylation. The dephosphorylated form does not bind to DEAE-cellulose. Inorganic phosphate and serine phosphate but not mannose 6-phosphate can inhibit this dephosphorylation. The kinetic properties of the phosphorylated and dephosphorylated arylsulfohydrolase are quite similar. The possibility that arylsulfohydrolase B is a dephosphorylated form of B, has been ruled out by the significant differences between substrate concentration and activity curves of these enzymes. 

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