AbstractsBiology & Animal Science

The human genome expresses 6 proteins of the family 18 chitinases, whereof YKL-40 (CHI3L1) is the most extensively studied. This protein binds chitin-derived oligosaccharides (ChOS) with micromolar affinity, but lacks the ability to degrade chitin. Besides possessing a chitin binding site, YKL-40 is also known to bind to heparin and collagen. Knowledge on the biological role of YKL-40 is still superficial but its function has mostly been related to immune signaling, embryonic development and remodeling of tissues and extracellular matrixes. This is particularly relevant in pathological conditions involving inflammation and connective tissue remodeling; such as in rheumatoid arthritis, osteoarthritis, liver fibrosis and many cancer diseases. Severity of the disease is usually directly linked to increased serum concentration of YKL-40. Currently, the focus in YKL-40 research is gradually shifting from diagnostic marker to a putative therapeutic target in various diseases. For a better understanding of the physiological activity and the underlying biochemical mechanisms, it is of great interest to examine the binding affinity of YKL-40 to its putative binding ligands. In this study, methods for maximal expression of histidine tagged YKL-40 protein were optimized using his-tagged YKL-40 transfected HEK293 cells in an adherent cell culture. A downstream purification process was tested using nickel and heparin columns. The yield of purified YKL-40 was not in accordance with the amount of YKL-40 in the cell medium, as determined by ELISA. However, eliminating fetal bovine serum (FBS) from the cell medium improved the yield dramatically. Hence, FBS was concluded to contain components interfering with the purification process; only YKL-40 obtained from FBS free cell cultures could be used for successful purification. The purified YKL-40 was applied in YKL-40 – heparin binding studies, where the protein was found to bind firmly and quantitatively to an immobilized heparin resin. Erfðamengi mannsins tjáir sex prótein sem tilheyra kítínasafjölskyldu 18, en af þeim hefur YKL-40 (CHI3L1) mest verið rannsakað. Prótein þetta bindur fásykrur úr kítíni (ChOS) með míkrómólar bindisækni, en getur ekki brotið kítín niður. Auk bindisets fyrir kítín er vitað að YKL-40 getur einnig bundið heparín og kollagen. Um líffræðilegt hlutverk YKL-40 er enn lítið vitað en helst hefur verið talið að starf þess tengist boðmiðlun ónæmiskerfisins, fósturþroskun og ummyndun vefja og millifrumuefnis. Þetta skiptir sérstaklega máli í sjúkdómsástandi þar sem bólgusvörun og ummyndun bandvefs á sér stað, svo sem í iktsýki, slitgigt, skorpulifur og mörgum sjúkleika af völdum krabbameins. Að jafnaði er alvarleiki sjúkdómsins beintengdur hækkuðum styrk YKL-40 í serumi. Um þessar mundir beinast rannsóknir á YKL-40 síður að því sem sjúkdómsgreiningarmerki og frekar að því sem meintu viðfangi til lækninga ýmissa sjúkdóma. Til að auka skilning á lífeðlisfræði próteinsins og lífefnafræðilegum ferlum sem að baki búa er áhugavert að rannsaka hæfni YKL-40 til að…