Skalp/ elafin: molecular cloning, expression, purification and characterization as immunomodulator.

TATEOSIAN N; COSTA J; MAFIA P; GUERRIERI D; CHULUYAN E.

Cordoba, Argentina

Congreso; 7º congreso de la Asociacion Latinoamericana de Inmunologia; 2005

ALAI

AbstractSKALP/ ELAFIN (skin derived anti leukoproteinase) is a 9.9kDa serine proteinase inhibitor which contain multiple transglutaminase substrate domains. The aim of the present work was to clone and characterize their function on immune cells. A cDNA fragment coding for human SKALP/ ELAFIN was obtained by RT PCR from Hela cell line, and cloned into expression vector (pTYB2). This vector utilizes the inducible self-cleavage activity of intein (a protein splicing element) to isolate the target protein from the affinity tag. This construction was used to transform E. Coli strain (ER2566) competent cells. Clones were screened by colony PCR and sequenced. Once the protein was isolated and purified, the elastase inhibitory activity of rhSKALP/ ELAFIN was measured with a chromogenic substrate. We next examine the effect of rhSKALP/ ELAFIN on OKT3 and IL-2 induced-lymphocyte proliferation. Like other serpins, rhSKALP/ELAFIN was able to decrease the lymphocyte proliferation (table), and this was not by the induction of apoptosis. Furthermore, rhSKALP/ ELAFIN also decreased the expression of CD25 on CD4 T cells. Overall, these results suggest that rhSKALP/ELAFIN is an immunomodulator by decreasing lymphocyte proliferation. SKALP/ELAFIN (mg/ml) OKT3 IL-2 P < 0.0 41.0 ± 1.6 20.0 ± 4.0   0.4 11.6 ± 2.4   0.01 2.0 5.4 ± 1.5   0.01 4.0 2.7 ± 1.8 4.9 ± 1.6 0.01