다발구멍장이버섯으로부터 분리한 혈전용해 세린분해효소의 특성 연구

Abstract

3단계를 거처 식용 가능한 다발구멍장이버섯으로부터 분리한 혈전용해효소의 비활성은 30.49 U/mg이었으며, MALDI-TOF/MS로부터 분자량이 30086.41 Da이었다. 일차 아미노산 구조는 Glu-Thr-Val-Thr-Glu-Thr-Thr-Ala-Pro-Trp-Gly-Leu-Ser-Arg-Ile으로 밝혀졌으며, pH 8.0에서 pH 10.0의 넓은 범위에서 큰 활성을 나타내는 alkaline protease로, 최적온도는 <TEX>$50^{\circ}C$</TEX>이며, <TEX>$30^{\circ}C$</TEX>까지는 대체로 열에 안정한 효소였다. 이 효소는 합성 기질 N-Suc-Ala-Ala-Pro-Phe pNA을 강하게 분해하였으며, <TEX>$Hg^{2+}$</TEX> 금속이온의 첨가로 효소 활성이 완전히 사라졌다. 효소저해제인 PMSF의 첨가로 혈전용해활성이 사라지는 결과로부터 serine 분해효소임을 알 수 있었다. A fibrinolytic serine protease was purified from the fruiting bodies of an edible mushroom, Albatrellus confluens. The enzyme had a molecular mass of 30086.41 Da, as measured by MALDI-TOF mass spectrometry. The N-terminal amino acid sequence of the enzyme was Glu-Thr-Val-Thr-Glu-Thr-Thr-Ala -Pro-Trp-Gly-Leu-Ser-Arg-Ile. It displayed optimal activity at <TEX>$50^{\circ}C$</TEX> and within a pH range of <TEX>$8.0{\sim}10.0$</TEX>, suggesting that the enzyme is an alkaline protease. The enzyme was stable up to <TEX>$30^{\circ}C$</TEX>. The enzyme displayed a strong substrate specificity for the synthetic peptide, N-Suc-Ala-Ala-Pro-Phe pNA. The enzyme activity was completely inhibited by addition of PMSF, indicating that the enzyme is a serine protease. No inhibition was observed following addition of E-64, pepstatin, or EDTA. The activity of the purified enzyme was decreased in the presence <TEX>$Fe^{2+}$</TEX> or <TEX>$Co^{2+}$</TEX>, and the enzyme was completely inhibited by addition of <TEX>$Hg^{2+}$</TEX>. From these results, we propose that Albatrellus confluens could be used for biofunctional foods development and has potential therapeutic value for the treatment of vascular diseases.