Purification , Peptide Sequencing ,Characterisation and Study of

Antiproliferative activity of Cicer arietinum L. Protein

Phosphatase 2A on Chronic Myelogenous Leukemia K562 Cell line:

Ellucidation of Mechanism of Development of Chronic

Myelogenous Leukemia in Human Cell line.

Antik Kiron Bose Ph.D 1

1 Senior Scientist and Project Advisor, Human Biology Division, Fred Hutchinson Cancer Research

Center,Seattle, Washington.

Protein Phosphatase 2A has been purified from 72 hrs germinating

seeds of Cicer arietinum L. by Toyopearl DEAE -650 anion exchanger,

Phenyl-sepharose CL-4B ,Microcystin –aminoethanethiol- sepharose

chromatography ,PD-10 desalting column and Vydac C-18 Reverse phase

HPLC chromatography with 4186.67 folds of purification ,3% recovery

and final specific activity of 209.334 U/ �g protein /min .The molar

masses of catalytic subunit (38 KD) and regulatory subunits (65 and 75

KD) were determined by 6% SDS-PAGE .38 KD catalytic subunit was

identified by Western Blotting with anti IgG2 monoclonal Ab against

human PP2A catalytic subunit.Number of SH groups was found to be 11

in PP2A/c and its full peptide sequencing gave a 306 aa protein which

was deposited in Protein Public database Uniprot with Acc no. Q07098

.Human PP2A was found to be inhibited in human CML cell line K562 due

to Bcr:abl induced formation of binary complex between human PP2A/c

and hnRNP A2 and ternary complex between human PP2A holoenzyme,hn

RNP A2 and SET protein .In K562 mutant OA200 cell line lacking human

PP2A ; chick pea PP2A was found to induce apoptosis of CML cells as it

is not inhibited by hn RNP A2 or SET proteins in vivo or in

vitro.Chick pea PP2A further induced nuclear localization of

SET-NUP214 complex and degradation of bcr:abl fusion proteins .RNA

binding domains of hn RNP A2 was sufficient to inhibit human PP2A .

Chick pea PP2A associated with human PR72 and induced

dephosphorylation of SV 40 T antigen. pH and temperature optima of

chick pea PP2A were 8.0 and 25 �c respectively and Km and V max were

87.713 mM , 166.67 U/min for holoenzyme and 64.10 mM and 208.33 U/min

for PP2A respectively.