title: A Combined Molecular Cloning and Mass Spectrometric Method to Identify, Characterize, and Design Frenatin Peptides from the Skin Secretion of Litoria infrafrenata.
abstract: Amphibian skin secretions are unique sources of bioactive molecules, particularly bioactive peptides. In this study, the skin secretion of the white-lipped tree frog (Litoria infrafrenata) was obtained to identify peptides with putative therapeutic potential. By utilizing skin secretion-derived mRNA, a cDNA library was constructed, a frenatin gene was cloned and its encoded peptides were deduced and confirmed using RP-HPLC, MALDI-TOF and MS/MS. The deduced peptides were identified as frenatin 4.1 (GFLEKLKTGAKDFASAFVNSIKGT) and a post-translationally modified peptide, frenatin 4.2 (GFLEKLKTGAKDFASAFVNSIK.NH₂). Antimicrobial activity of the peptides was assessed by determining their minimal inhibitory concentrations (MICs) using standard model microorganisms. Through studying structure-activity relationships, analogues of the two peptides were designed, resulting in synthesis of frenatin 4.1a (GFLEKLKKGAKDFASALVNSIKGT) and frenatin 4.2a (GFLLKLKLGAKLFASAFVNSIK.NH₂). Both analogues exhibited improved antimicrobial activities, especially frenatin 4.2a, which displayed significant enhancement of broad spectrum antimicrobial efficiency. The peptide modifications applied in this study, may provide new ideas for the generation of leads for the design of antimicrobial peptides with therapeutic applications.
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. dwu03@qub.ac.uk.
authors: Di
authors: D
authors: Wu
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. ygao07@qub.ac.uk.
authors: Yitian
authors: Y
authors: Gao
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. l.wang@qub.ac.uk.
authors: Lei
authors: L
authors: Wang
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. x.xi@qub.ac.uk.
authors: Xinping
authors: X
authors: Xi
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. ywu16@qub.ac.uk.
authors: Yue
authors: Y
authors: Wu
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. m.zhou@qub.ac.uk.
authors: Mei
authors: M
authors: Zhou
authors: Department of Emergency Medicine, The First Hospital of Hebei Medical University, Shijiazhuang 050031, China. zhangyingqi08@sina.com.
authors: Yingqi
authors: Y
authors: Zhang
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. c.ma@qub.ac.uk.
authors: Chengbang
authors: C
authors: Ma
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. t.chen@qub.ac.uk.
authors: Tianbao
authors: T
authors: Chen
authors: Natural Drug Discovery Group, School of Pharmacy, Queen's University, Belfast BT9 7BL, Northern Ireland, UK. chris.shaw@qub.ac.uk.
authors: Chris
authors: C
authors: Shaw
mesh_terms: D000595:Amino Acid Sequence; D029845:Amphibian Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D000890:Anti-Infective Agents / Q000138:chemical synthesis / Q000494:pharmacology; D001001:Anura / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D015195:Drug Design; D008826:Microbial Sensitivity Tests; D010455:Peptides / Q000138:chemical synthesis* / Q000235:genetics / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational; D012867:Skin / Q000378:metabolism*; D013329:Structure-Activity Relationship; D053719:Tandem Mass Spectrometry
chemical_list: D029845:Amphibian Proteins; D000890:Anti-Infective Agents; D010455:Peptides
keywords: antimicrobial peptide; frenatin; frog skin secretion; modification; structure activity relationship