DNA-Polymerases α, δ and ε from T-Cell Spontaneous Lymphoblastic Leukemia of Sprague-Dawley Inbred Rat: Isolation and Characterization

[img]PDF - Authorized users only
Language: English
197Kb
Title:DNA-Polymerases α, δ and ε from T-Cell Spontaneous Lymphoblastic Leukemia of Sprague-Dawley Inbred Rat: Isolation and Characterization
Creators:
Kramata, Pavel
Černý, Jaroslav
Birkuš, Gabriel
Votruba, Ivan
Otová, Berta
Holý, Antonín
Journal or Publication Title:
Collection of Czechoslovak Chemical Communications, 60, 9, pp. 1555-1572

Abstract

<p>Using a single isolation procedure and selective assays for the determination of enzyme activity, highly purified DNA-polymerases α, δ and ε were isolated from the lymphoma of Sprague-Dawley inbred rats. For pol α the subunit composition was 170, 70, 57 and 53 kDa with sedimentation coefficient 8.7 S for the native molecule; pol delta consists of two polypeptides (133 and 46 kDa; sedimentation coefficient 8.2 S), while pol ε is a single polypeptide (140 kDa) and its sedimentation coefficient is 7.0 S. Comparison of the interaction of individual enzymes with known inhibitors and proliferating cell nuclear antigen (PCNA) using the template-primer poly dA-oligo dT<sub>12-18</sub>, gave the following data: (i) pol α is selectively inhibited by N<sup>2</sup>-(p-butylphenyl)-2'-deoxyguanosine 5'-triphosphate (BuPdGTP) and stimulated by dimethyl sulfoxide; (ii) all the enzymes are inhibited by N-ethylmaleimide and aphidicolin; (iii) PCNA stimulates pol δ approximately 50 times while pol ε is moderately inhibited; (iv) pol α exhibits considerably higher DNA-primase activity with poly dC as template than with poly dT, and negligible 3'-5'-exonuclease activity whereas pol δ and pol ε, which do not exert any DNA-primase activity have approximately the same 3'-5'-exonuclease activity. The ability of individual polymerases to utilize poly dT-oligo dA<sub>12-18 </sub>as a template-primer at different pH values, ionic strengths and Mg<sup>2+</sup>-concentrations was also investigated. In comparison to poly dA-oligo dT<sub>12-18</sub> template-primer, pol α has 140% of enzyme activity on poly dT-oligo dA<sub>12-18</sub> under optimal conditions, whereas the activity of pol ε and pol δ is 4 times and 10 times lower, respectively.

Title:DNA-Polymerases α, δ and ε from T-Cell Spontaneous Lymphoblastic Leukemia of Sprague-Dawley Inbred Rat: Isolation and Characterization
Creators:
Kramata, Pavel
Černý, Jaroslav
Birkuš, Gabriel
Votruba, Ivan
Otová, Berta
Holý, Antonín
Divisions:Life and Chemical Sciences > Institute of Organic Chemistry and Biochemistry > Collection of Czechoslovak Chemical Communications
Journal or Publication Title:Collection of Czechoslovak Chemical Communications
Volume:60
Number:9
Page Range:pp. 1555-1572
ISSN:0010-0765
E-ISSN:1212-6950
Publisher:Institute of Organic Chemistry and Biochemistry
Related URLs:
URLURL Type
http://dx.doi.org/10.1135/cccc19951555UNSPECIFIED
ID Code:852
Item Type:Article
Deposited On:06 Feb 2009 17:04
Last Modified:06 Feb 2009 16:04

Citation

Kramata, Pavel; Černý, Jaroslav; Birkuš, Gabriel; Votruba, Ivan; Otová, Berta; Holý, Antonín (1995) DNA-Polymerases α, δ and ε from T-Cell Spontaneous Lymphoblastic Leukemia of Sprague-Dawley Inbred Rat: Isolation and Characterization. Collection of Czechoslovak Chemical Communications, 60 (9). pp. 1555-1572. ISSN 0010-0765

Repository Staff Only: item control page