terminal transferase
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2020 ◽  
Vol 56 (63) ◽  
pp. 8912-8915
Author(s):  
Hyo Yong Kim ◽  
Jayeon Song ◽  
Ki Soo Park ◽  
Hyun Gyu Park

A personal glucose meter-based terminal transferase activity assay utilizing the glucose oxidase-mimicking activity of cerium oxide nanoparticles was developed.


2019 ◽  
Vol 47 (13) ◽  
pp. 6932-6945 ◽  
Author(s):  
Ankita Gupta ◽  
Shailesh B Lad ◽  
Pratibha P Ghodke ◽  
P I Pradeepkumar ◽  
Kiran Kondabagil

Abstract Acanthamoeba polyphaga mimivirus is an amoeba-infecting giant virus with over 1000 genes including several involved in DNA replication and repair. Here, we report the biochemical characterization of gene product 577 (gp577), a hypothetical protein (product of L537 gene) encoded by mimivirus. Sequence analysis and phylogeny suggested gp577 to be a primase-polymerase (PrimPol)—the first PrimPol to be identified in a nucleocytoplasmic large DNA virus (NCLDV). Recombinant gp577 protein purified as a homodimer and exhibited de novo RNA as well as DNA synthesis on circular and linear single-stranded DNA templates. Further, gp577 extends a DNA/RNA primer annealed to a DNA or RNA template using deoxyribonucleoties (dNTPs) or ribonucleotides (NTPs) demonstrating its DNA/RNA polymerase and reverse transcriptase activity. We also show that gp577 possesses terminal transferase activity and is capable of extending ssDNA and dsDNA with NTPs and dNTPs. Mutation of the conserved primase motif residues of gp577 resulted in the loss of primase, polymerase, reverse transcriptase and terminal transferase activities. Additionally, we show that gp577 possesses translesion synthesis (TLS) activity. Mimiviral gp577 represents the first protein from an NCLDV endowed with primase, polymerase, reverse transcriptase, terminal transferase and TLS activities.


2019 ◽  
Vol 47 (1) ◽  
pp. 256-259 ◽  
Author(s):  
Bhagwan S. Batule ◽  
Chang Yeol Lee ◽  
Ki Soo Park ◽  
Hyun Gyu Park

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 5194-5194
Author(s):  
Aldair Sousa Paiva ◽  
Hugo Diogenes De Oliveira Paiva ◽  
Geraldo Barroso Cavalcanti ◽  
Frank Bahia ◽  
Rodrigo Villar Freitas ◽  
...  

Abstract Background: The detection of Intracellular (IC) antigens by flow cytometry (FC) such as myeloperoxidase (MPO), cCD13, cCD79a, cCD22, cCD3 and Terminal deoxynucleotidyl Transferase (TdT) has become the useful tool in the differential diagnosis between acute myeloid leukemias (AML) and acute lymphoid leukemias (ALL). Through detection of myeloid antigens (MPO and cCD13), B cells precursors (cCD79a and cCD22) and precocity T-cells (cCD3) it has been possible to confirm the diagnosis of these acute leukemias. The detection of intracellular cell markers by FC usually requires previous permeabilization of fresh cell suspensions. TdT, also known as DNA nucleotidylexotransferase (DNTT) or terminal transferase, is a specialized DNA polymerase expressed in immature, pre-B, pre-T lymphoid cells, and acute lymphoblastic leukemia/lymphoma cells. TdT adds N-nucleotides to the V, D, and J exons of the TCR and BCR genes during antibody gene recombination, enabling the phenomenon of junctional diversity. In humans, terminal transferase is encoded by the DNTT gene. This antigen is expressed mostly in the nucleus cells from primary lymphoid organs, like the thymus and bone marrow. The TdT detection has also been shown to be useful in confirming the acute forms of B and T-lineage lymphoproliferative diseases by FC. The aim of this study was to demonstrate the importance of this cell markers' detection by FC in the differential diagnostic of acute leukemias. Methods: Bone marrow and/or peripheral blood leukemic cells from 50 cases of acute leukemia: 16 ALL and 36 AML. The cells were fixed and permeabilized in briefly exposed to Becton & Dickinson Lyse Solution at concentration of 10%, and subsequently labeled with monoclonal antibodies anti-MPO, TdT, CD3, CD13, CD22 and CD79a. Results: The MPO expression was observed in 35/36(97,22%) and cCD13 in all cases of AML and in none ALL patients. Three cases of MPO-positive ALL (FAB-L2) could be reclassified as M0-AML. These cases were CD34+;HLADR+;CD33-;CD13-;CD7+ and cCD13+. The intensity of TdT expression was observed in 15/16 (93.8%) of ALL and 5/36 (13.9%) of AML. The cCD22 and cCD79a were positive in 15/16 (93.8%) and all of pre-B ALL respectively and cCD3 was expressed in one case of Pre-T ALL that initial phenotype was CD34+/HLADR+/TdT+/CD7+ and sCD3-). Conclusions: These results indicate that monoclonal antibodies anti-MPO, cCD13, cCD79a, cCD22, cCD3 and TdT were excellent cell markers for the diagnosis and classification of acute leukemias and can be reliably detected by FC. This rapid and specific technique should be a valuable addition to routine immunophenotyping of acute leukemia. Disclosures No relevant conflicts of interest to declare.


eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Tatiana Kent ◽  
Pedro A Mateos-Gomez ◽  
Agnel Sfeir ◽  
Richard T Pomerantz

DNA polymerase θ (Polθ) promotes insertion mutations during alternative end-joining (alt-EJ) by an unknown mechanism. Here, we discover that mammalian Polθ transfers nucleotides to the 3’ terminus of DNA during alt-EJ in vitro and in vivo by oscillating between three different modes of terminal transferase activity: non-templated extension, templated extension in cis, and templated extension in trans. This switching mechanism requires manganese as a co-factor for Polθ template-independent activity and allows for random combinations of templated and non-templated nucleotide insertions. We further find that Polθ terminal transferase activity is most efficient on DNA containing 3’ overhangs, is facilitated by an insertion loop and conserved residues that hold the 3’ primer terminus, and is surprisingly more proficient than terminal deoxynucleotidyl transferase. In summary, this report identifies an unprecedented switching mechanism used by Polθ to generate genetic diversity during alt-EJ and characterizes Polθ as among the most proficient terminal transferases known.


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