鶏血漿中 Aspartate-および Alanine-aminotransferase 系アミノ基転移酵素の分別法について

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As was reported previously31=34?36), aspartate aminotransferase (GOT) and alanineaminotransferase (GPT) in fowl blood plasma can be classified roughly into four typesunder the general terms of forward(F)-GOT, reverse (R)-GOT, F-GPT, and R-GPT.The author36) succeeded considerably in overcoming the difficulties of fractionatingthese transaminases by using an ingenious method introduced by COHNIO) and 0NCLEY25).Some of the transaminases fractionated were rich in lipids and contained such proteincomponents of particular kind as beta.-metal combining protein28,35) and heme prote-j.35)In order fully to purify these transaminases, it was necessary to remove lipids andparticular components. For the purpose of meeting this necessity, the author devised amethod. The following diagram is a schematic illustration of this method for separationof the four fractions involved in the transamination reaction.A.B.C.Removal of lipids from fowl blood plasma with n-butanol.To I volume of plasma was added I volume of n-butanol. A mixture wassturred at 0C [01 30 II11I1UtCS and centrufuged It 700 X g at OC for 30 minutes.The supernatant and the insoluble middle layer were discarded. The solublelowermost layer at pH 8.4 was saved as source sample I.Fractional precipitation with ammonium sulfate.Source sample I was used for a partial fractionation of the transaminases bystepwise additions of ammonium sulfate. Some of the filtrates were pooledas source sample 2.In addition, salting out was performed by Osbornes method.Sephadex get fractionation of source sample 2.Sourse sample 2 was get filtrated through Sephadex G-100 and with phosphatebuffer at pH 7.4 as eluant.Some of the filtrates were pooled as source sample 3. Samples were usually maintained at 0C throughout an experiment for purification.The results obtained are summarized as follows.l. It was impossible to recover fractiorns I, [14111, and IV-I from source sample 3by the 6th method of COIN. These fractions could be removed by means of a methoddevised by the author (n-butanol extraction, salting out, and Sephadex get fractionation).Source sample 3 also contained fractions IV-4, V, and V-l.2. Fraction IV-5 was separated by the authors method. It was useful for studieson the possible role of lipoprotein in the transaminatioru reaction of particular kind.3. There was 110 good agreement among the transaminases in the mode of fractionalprecipitation. R-GOT and R-GPT, however, agreed verv well with each other in thismode and partially with F-GPT. It is of interest to speculate from these results thatF-GPT, R-GOT, and R-GPT may have a common ancestry. To lend support to thisspeculation, discussion will be made in a future report on the possible effect of fractionIV-8 on the reversible transamination reaction of GPT.From the results described above, it seems possible to reocver this reversible GPT.It is clear that there is no correlation betweern F-GOT and any other transaminase.
社団法人日本獣医学会の論文
1972-08-25

鶏血漿中 Aspartate-および Alanine-aminotransferase 系アミノ基転移酵素の分別法について

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