枯草桿菌中的胞外金屬蛋白酶 (Extracellular metalloprotease from Bacillus subtilis, Mpr；EC 3.4.21.19 ) 屬於serine peptidases中的glutamyl endopeptidase (clan PA family S1 subfamily A，S1A) 。枯草桿菌中的Mpr擁有narrow substrate specificity，具專一性水解glutamate及aspartate中的□-carboxyl groups之能力，可以應用在特定融合蛋白質之分解，進而產生生物活性胜肽 (bioactive peptides) 。生物活性胜肽已知對人體的生理功能具有正面的影響。然而，在自然界中生物活性胜肽的含量通常不多，因此有許多研究都致力於增加食物中生物活性胜肽的含量。為了達到這個目標，發展出具有受質專一性的蛋白酶是必要的。本論文主要目的為決定Mpr的X光三度空間結構，以進行後續酵素工程之研究。我們利用overlapping PCR方法，得到pro-Mpr S267A突變株，並將其表現的蛋白質形成晶體使用多波長異常散射方法 (multi-wavelength anomalous dispersion methods, MAD) 得到解析度為1.1 Å的pro-Mpr S267A 突變株的結構。如同其他的S1A family chymotrypsin-like fold蛋白酶，整個pro-Mpr S267A突變株的結構由N-domain和C-domain組成，這兩個domain分別由六個和七個反向平行的□-sheets形成□-barrels。所形成的□-barrels會相互垂直，且在這兩個domain之間形成一個具有酵素催化位置及受質結合位置的夾縫。pro-Mpr S267A 突變株擁有的propeptide區域環繞在mature-Mpr的外圍，提供適當的骨架使mature-Mpr能摺疊成正確的構型。此外，propeptide片段 (residue 85-93) 會遮蔽Mpr的催化位置及受質結合位置，推測在有propeptide存在時，Mpr無法進行其酵素功能。藉由pro-Mpr S267A突變株的3D結構所提供的資訊，可並進一步修飾其受質結合位期望能增加酵素的受質專一性。未來工作將以結構為基礎，探討其受質結合位置的相關性，並進一步修飾其受質結合位置，期望能增加酵素的受質專一性，對於提昇合成生物活性的蛋白質與胜肽的商業化應用將有相當大的助力。

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