Determination of potential glycosylation profile for human telomerase's htert subunit in hepatocellular carcinoma and neuroblastoma cell line with bioinformatic and experimental methods

Telomerase, an eukaryotic ribonucleoprotein, is a holoenzyme complex consisting of proteins linked together such as "HTERT" catalytic subunit, "hTR" template RNA and "p23 / hsp90", "SnoRNA", "TEP1", "hNRNAPS C1 / 2 "," La "," L22 "and" hStau. At each cell division, the ends of the chromosomes (telomeres) are shortened. With this shortening, the stopping of cell senescence and cell division is triggered. Telomerase is a reverse transcriptase that catalyzes the insertion of new nucleotides into telomeres (human: TTAGGG nucleotides) and has been identified in about 90% of all malignant tumors. When human tumor cells are thought to bypass the global pathways of cellular senescence and DNA damage signaling. So, telomerase is at a critical point in terms of the relevant molecular mechanisms. Protein glycosylation, a post-translational modification, has a number of important molecular roles, such as intracellular and intercellular recognition, adhesion, signal transduction and differentiation. Protein glycosylation is particularly important in the large proportion of biological recognition events, such as quality control of the binding and protein folding. From the perspective of glycobiology, telomerase, a multi-subunited ribonucleoprotein; it is possible that glycans play a role in the telomerase's subunits' form the telomerase complex and the necessary recognition and attachment reactions of the relevant molecular mechanisms of the catalytic function of the enzyme. In this thesis study, it is primarily aimed to determine possible glycosylation positions in the amino acid sequences of hTERT and telomerase catalytic subunit (TERT) that belongs to different species, by bioinformatic methods; and to experimentally compare and analyze possible glycans from Neuroblastoma (SH- SY5Y) and Hepatocellular carcinoma (Hep G2) cell line samples by western and lectin blotting methods. As a result of the bioinformatics study, it was found that when comparing the TERT protein in different species with respect to the amino acid sequence, these sequences are highly conserved between species and differ in terms of folding motifs in the protein secondary models. In the amino acid sequence of the Homo sapiens hTERT, an N-linked and multiple O-linked glycosylation positions were identified. In addition, possible O-linked glycosylation positions have been identified in the TERT amino acid sequences of Canis familiaris, Bos taurus, Rattus norvegicus and Mus musculus and two possible N-linked glycosylation positions were found in the TERT amino acid sequences of Rattus norvegicus and Mus musculus. In support of bioinformatics data, N-acetylglucosamine (GlcNAc) and sialic acid (Sia) bound to hTERT were identified in the nucleus localization of SH-SY5Y and Hep G2 cells. GlcNAc which is bound to hTERT has been found most intensively in both cell lines. Additionally hTERT-associated mannose was detected in Hep G2 cells. The α2,6-linked Sia, identified as end-sugar in hTERT of both cell types, might be associated with O- and/or N-linked glycosylation; and it is discussed that these α2,6-linked Sias might have attachment and recognition effects on molecular reactions of hTERT, and an interacting protein TEP-1's binding Key words: Telomerase; hTERT; Glycosylation; Neuroblastoma; Hepatocellular carcinoma, Sialic acid, GlcNAc, Mannose, Western/Lectin blotting