title: Facile expression and purification of the antimicrobial peptide histatin 1 with a cleavable self-aggregating tag (cSAT) in Escherichia coli.
abstract: Human histatin 1 (Hst1), a member of the histatin family, possesses antimicrobial properties. In this study, we applied a previously developed cleavable self-aggregating tag (cSAT) for the expression and purification of histatin 1 to demonstrate its utility for peptide expression and purification. The tag consists of a self-cleavable intein and a self-assembling peptide ELK16 (I-ELK16). First, an active insoluble aggregate of the recombinant histatin 1-Mxe GyrA intein-ELK16 (Hst1-I-ELK16) fusion protein was produced with a yield of 28.9 μg/mg wet cell pellet. The thiol reagent dithiothreitol (DTT) was then used to induce the intein-mediated cleavage and peptide release into the soluble fraction with a yield of 2.06 μg/mg wet cell pellet and a purity of 70%. The peptide was further purified by high performance liquid chromatography. These results were comparable to the yield and purity achieved when the more conventional glutathione transferase (GST) tag was used. The antimicrobial activities of this recombinant histatin 1 were confirmed against three Candida strains. This cSAT technique offers considerable advantages in terms of its simplicity and speed, eliminating the need for an exogenous protease, and reducing the number of chromatography purification steps. This technique should also be useful for the expression and purification of other AMPs.
authors: Department of Chemical Engineering, Tsinghua University, One Tsinghua Garden Road, Beijing 100084, China. xingl08@mails.tsinghua.edu.cn
authors: Lei
authors: L
authors: Xing
authors: Wanghui
authors: W
authors: Xu
authors: Bihong
authors: B
authors: Zhou
authors: Yilu
authors: Y
authors: Chen
authors: Zhanglin
authors: Z
authors: Lin
mesh_terms: D000595:Amino Acid Sequence; D000890:Anti-Infective Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism* / Q000494:pharmacology; D002175:Candida / Q000187:drug effects; D002177:Candidiasis / Q000188:drug therapy; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D054808:Histatins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification* / Q000494:pharmacology; D006801:Humans; D047668:Inteins; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000890:Anti-Infective Agents; D054808:Histatins; D011993:Recombinant Fusion Proteins; C068074:HTN1 protein, human