title: Exoproteome of Aspergillus flavus corneal isolates and saprophytes: identification of proteoforms of an oversecreted alkaline protease.
abstract: Aspergillus flavus infects the human eye leading to keratitis. Extracellular proteins, the earliest proteins that come in contact with the host and virulence related exoproteins, were identified in the fungus isolated from infected cornea. Virulence of the corneal isolates was tested in the Galleria mellonella larvae model and those isolates showing higher virulence were taken for subsequent exoproteome analysis. High resolution two-dimensional electrophoresis and mass spectrometry were used to generate A. flavus exoproteome reference map as well as to profile most of the exoproteins. Analysis of the identified proteins clearly shows the major biological processes that they are involved in. Nearly 50% of the exoproteins possess catalytic activity and one of these, an alkaline serine protease (Alp1) is present in high abundance as well as multiple proteoforms. Many proteins in the A. flavus exoproteome have been shown to be virulence factors in other pathogens indicating the probable role for these proteins in the corneal infection as well. Interestingly, the majority of the exoproteins do not have secretory signal indicating that they are secreted through the non-classical pathway. Thus, this study provides a clue to the early strategies employed by the pathogen to establish an infection in an immunocompetent host.
  
  
  CONCLUSIONS
  The outcome of a fungal infection in an immunocompetent human eye depends on the ability of the fungus to overcome the host defense and propagate itself. In this process, the earliest events with respect to the fungal proteins involved include the secretory proteins of the invading organism. As a first step towards understanding the role of the extracellular proteins, exoproteome profile of the fungal isolates was generated. The fungal isolates from cornea showed a distinct pattern of the exoproteome when compared to the saprophyte. Since corneal isolates also showed higher virulence in the insect larval model, presumably the proteins elaborated by the corneal isolates are virulence related. One of the abundant proteins is an alkaline serine protease and this protein exists as multiple proteoforms. This study reports the comprehensive profile of exoproteome and reveals proteins that are potential virulence factors.
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Ramu Muthu
authors.initials: RM
authors.lastname: Selvam
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Rathnavel
authors.initials: R
authors.lastname: Nithya
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Palraj Narmatha
authors.initials: PN
authors.lastname: Devi
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: R S Bhuvana
authors.initials: RS
authors.lastname: Shree
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Murugesan Valar
authors.initials: MV
authors.lastname: Nila
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Naveen Luke
authors.initials: NL
authors.lastname: Demonte
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Chitra
authors.initials: C
authors.lastname: Thangavel
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Jayapal Jeya
authors.initials: JJ
authors.lastname: Maheshwari
authors.affiliation: Department Of Ocular Microbiology, Aravind Eye Hospital, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Prajna
authors.initials: P
authors.lastname: Lalitha
authors.affiliation: Cornea Clinic, Aravind Eye Hospital, Aravind Eye Care System, Madurai, Tamil Nadu, India.
authors.forename: Namperumalsamy Venkatesh
authors.initials: NV
authors.lastname: Prajna
authors.affiliation: Proteomics Department, Aravind Medical Research Foundation, Dr. G. Venkataswamy Eye Research Institute, Aravind Eye Care System, Madurai, Tamil Nadu, India. Electronic address: dharmalingam.k@gmail.com.
authors.forename: Kuppamuthu
authors.initials: K
authors.lastname: Dharmalingam
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism*; D000818:Animals; D001228:Aspergillosis / Q000378:metabolism*; D001231:Aspergillus flavus / Q000302:isolation & purification / Q000378:metabolism / Q000472:pathogenicity; D003315:Cornea / Q000382:microbiology; D003316:Corneal Diseases / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology; D004195:Disease Models, Animal; D005656:Fungal Proteins; D006801:Humans; D009036:Moths; D020543:Proteome / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D020543:Proteome; D000469:Alkaline Phosphatase
keywords: Aspergillus flavus; Exoproteome; Galleria mellonella; Mass spectrometry; Mycotic keratitis