Abstract
有些熱休克蛋白質可能具有分子監護子功能,而有些熱休克蛋白質其功能類似Caseinolytic protease (Clp)的蛋白質,在高溫逆境時能降解已受損害的蛋白質.Clp是一高度保守的蛋白質家族所組成,能存在於所有的生物體中.其中之ClpA明顯地存在於所有的真核生物體與原核生物體中,六個單體的ClpA會和十二個單體的ClpP形成一個依賴於ATP的Clp 蛋白酵素多聚體,能水解異常蛋白質.處於逆境下,ClpP-ClpA蛋白酵素便可能降解變性的蛋白質,藉此幫助細胞繼續存活於不利的環境.本實驗之目的是為探討阿拉伯芥之clpA-like基因的結構及其表現與高溫逆境的關係.以豌豆clp cDNA(Pspclp)做探針,從阿拉伯芥的cDNA基因庫中篩選出一段clpA-like基因,並將此cDNAp嵌入Bluescript II KR+質體,定名為AtclpaA.AtclpA全長為2244個鹼基對,可能缺少部份5'序列,其不完全的1998個鹼基對開放讀碼可轉譯成666個胺基酸,並含3'端208個開放讀碼的鹼基對,及長度為38個鹼基對的poly (A)尾巴,其開放讀碼區域與Brassica napus的clpA-like 基因(BnclpA)相似度很高.分析阿拉伯芥的基因組結果顯示AtclpA基因可能係以低拷貝數存在於植物中,亦可能含有內含子.AtclpA mRNA之表現不因高溫(37度)處理而增加.這些實驗結果提供進一步探討AtclpA在阿拉伯芥的表現及推測基因產物之基本資料.Several Hsps may function as molecular chaperones and some Hspsmay promote degradation of heat-damaged proteins during heatstress.Caseinolytic protease-like (Clp-like) proteins comprise ahightly conserved protein family, members of which have beenidentified in all organisms.ClpA and ClpB form a complex whichis an ATP-dependent protease composed of a dodecameric componentClpP and a hexameric component ClpA.The processive degradationof denatured proteins by ClpP-ClpA complex protects cells tosurvive in the detrimental environment.Our objective was tostudy the struture of the Arabidopsis clp-lke gene (AtclpA) andthe relationships between its expression and heat stress.In thisstudy, a clpA-like gene was isolated from an Arabidopsis cDNAlibrary using the pea clp cDNA (Pspclp) as a probe.TheArabidopsis clpA-like gene was subcloned into pBluescript II KS+vector and named AtclpA.AtclpA cDNA is 2244 nucleotides inlength consisting of a 1998-nucleotide incomplete open readingframe with a 208-nucleotide 3' untranslated sequence plus apoly(A) tail of 38 nucleotides.The open reading frame of theAtclpA shares 89.3% identity with BnclpA of Brassica napus.Analysis of genomic DNA indicates that AtclpA is present as alow-copy-number gene and may contain at least an intron.Hightemperature treature (37 degree) did not cause an increase anthe level of the AtclpA mRNA.Thesis results provide preliminaryinformation for further study on the expression, struture andfunction of the AtclpA.