經由生物資訊資料庫(National Center for Biotechnology Information，NCBI)之搜尋和比對方式找尋到一組在斑馬魚(編號AI477544)及人類(編號NP_115679)皆有表現之新穎17-beta羥基固醇去氫(17-beta Hydroxysteroid Dehydrogenase，17β HSD)基因。利用3’ Rapid Amplification of cDNA Ends(3’RACE)聚合連鎖反應的方法，以0~7天之斑馬魚胚胎之基因庫為模板，成功地放大並定序其核酸序列。此斑馬魚之開放閱讀框架(Open Reading Frame)具有1469個鹼基對，推定之胺基酸為416個;人類之開放閱讀框架為3003個鹼基對，345個胺基酸。此新穎之17-beta HSD具有短鏈酒精去氫(adh_short)與膽固醇攜帶蛋白(SCP2)二個酵素之結合結構區，因此假定此酵素之生理功能及酵素反應相似於固醇類物質之氧化還原。利用pQE表達系統，在大腸桿菌(E coli.)中表現及純化該斑馬魚及人類之新穎17-beta羥基固醇去氫(17β HSD)之蛋白質後，利用酵素動力學分析(enzyme kinetic assay)方法進行酵素反應之分析測試。結果發現此酵素以NAD+/NADP+為輔，Estriol和5-Androstene 3β,16α,17β -triol為受質時，可推動受質的氧化作用，因此該酵素可能是16α,17β dihydroxysteroid dehydrogenase。
In this study, by using BLAST (Basic Local Alignment Search Tool), a novel 17-beta Hydroxysteroid Dehydrogenase of zebra fish(accession number AI477544) and human(accession number NP_115679) were found from NCBI (National Center for Biotechnology Information) nucleotide database. The novel 17-beta Hydroxysteroid Dehydrogenase cDNA of zebra fish was successfully amplified from the 0-7 days old zebra fish embryo cDNA library by using the combination of PCR and 3’rapid-amplification of cDNA ends (3’RACE) methods. The novel zebra fish cDNA had a 1469 bp open reading frame and 416 deduced amino acid. The novel human cDNA had a 3303bp open reading frame and 345 amino acid. Both enzyme had two domain, i.e. short chain alcohol dehydrogenase(adh_short) and steroid carried protein(SCP2)domain. The initial velocity analysis of both enzyme showed that two substrates, Estriol and 5-Androstene 3β,16α,17β-triol may participate in the catalytic reaction of the novel 17-beta Hydroxysteroid Dehydrogenase.
