title: Molecular cloning and functional characterization of an antifungal PR-5 protein from Ocimum basilicum.
abstract: Pathogenesis-related (PR) proteins are involved in biotic and abiotic stress responses of plants and are grouped into 17 families (PR-1 to PR-17). PR-5 family includes proteins related to thaumatin and osmotin, with several members possessing antimicrobial properties. In this study, a PR-5 gene showing a high degree of homology with osmotin-like protein was isolated from sweet basil (Ocimum basilicum L.). A complete open reading frame consisting of 675 nucleotides, coding for a precursor protein, was obtained by PCR amplification. Based on sequence comparisons with tobacco osmotin and other osmotin-like proteins (OLPs), this protein was named ObOLP. The predicted mature protein is 225 amino acids in length and contains 16 cysteine residues that may potentially form eight disulfide bonds, a signature common to most PR-5 proteins. Among the various abiotic stress treatments tested, including high salt, mechanical wounding and exogenous phytohormone/elicitor treatments; methyl jasmonate (MeJA) and mechanical wounding significantly induced the expression of ObOLP gene. The coding sequence of ObOLP was cloned and expressed in a bacterial host resulting in a 25kDa recombinant-HIS tagged protein, displaying antifungal activity. The ObOLP protein sequence appears to contain an N-terminal signal peptide with signatures of secretory pathway. Further, our experimental data shows that ObOLP expression is regulated transcriptionally and in silico analysis suggests that it may be post-transcriptionally and post-translationally regulated through microRNAs and post-translational protein modifications, respectively. This study appears to be the first report of isolation and characterization of osmotin-like protein gene from O. basilicum.
authors.affiliation: CSIR-Indian Institute of Integrative Medicine (CSIR-IIIM), Canal Road, Jammu 180001, India.
authors.forename: Irshad Ahmad
authors.initials: IA
authors.lastname: Rather
authors.affiliation: CSIR-Indian Institute of Integrative Medicine (CSIR-IIIM), Canal Road, Jammu 180001, India.
authors.forename: Praveen
authors.initials: P
authors.lastname: Awasthi
authors.affiliation: CSIR-Indian Institute of Integrative Medicine (CSIR-IIIM), Canal Road, Jammu 180001, India; Academy of Scientific and Innovative Research, Council of Scientific & Industrial Research, Canal Road, Jammu 180001, India.
authors.forename: Vidushi
authors.initials: V
authors.lastname: Mahajan
authors.affiliation: CSIR-Indian Institute of Integrative Medicine (CSIR-IIIM), Canal Road, Jammu 180001, India; Academy of Scientific and Innovative Research, Council of Scientific & Industrial Research, Canal Road, Jammu 180001, India.
authors.forename: Yashbir S
authors.initials: YS
authors.lastname: Bedi
authors.affiliation: CSIR-Indian Institute of Integrative Medicine (CSIR-IIIM), Canal Road, Jammu 180001, India; Academy of Scientific and Innovative Research, Council of Scientific & Industrial Research, Canal Road, Jammu 180001, India.
authors.forename: Ram A
authors.initials: RA
authors.lastname: Vishwakarma
authors.affiliation: CSIR-Indian Institute of Integrative Medicine (CSIR-IIIM), Canal Road, Jammu 180001, India; Academy of Scientific and Innovative Research, Council of Scientific & Industrial Research, Canal Road, Jammu 180001, India. Electronic address: sumit@iiim.ac.in.
authors.forename: Sumit G
authors.initials: SG
authors.lastname: Gandhi
mesh_terms: D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D003001:Cloning, Molecular; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018649:Ocimum basilicum / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification* / Q000494:pharmacology; D016415:Sequence Alignment; D015003:Yeasts / Q000187:drug effects
chemical_list: D000935:Antifungal Agents; D010940:Plant Proteins
keywords: Antifungal; Elicitor; MicroRNA; Modeling; PR-5 protein; Thaumatin