The GPI Anchor Signal Sequence Dictates the Folding and Functionality of the Als5 Adhesin from Candida albicans

Mohammad Faiz Ahmad,Anwar Ali,Rohini Muthuswami,Sneha Sudha Komath,Samudrala Gourinath,Bhawna Yadav,Pravin Kumar,Amrita Puri,Mohit Mazumder,Scott G Filler

doi:10.1371/journal.pone.0035305

Abstract

BackgroundProteins destined to be Glycosylphosphatidylinositol (GPI) anchored are translocated into the ER lumen completely before the C-terminal GPI anchor attachment signal sequence (SS) is removed by the GPI-transamidase and replaced by a pre-formed GPI anchor precursor. Does the SS have a role in dictating the conformation and function of the protein as well?Methodology/Principal FindingsWe generated two variants of the Als5 protein without and with the SS in order to address the above question. Using a combination of biochemical and biophysical techniques, we show that in the case of Als5, an adhesin of C. albicans, the C-terminal deletion of 20 amino acids (SS) results in a significant alteration in conformation and function of the mature protein.Conclusions/SignificanceWe propose that the locking of the conformation of the precursor protein in an alternate conformation from that of the mature protein is one probable strategy employed by the cell to control the behaviour and function of proteins intended to be GPI anchored during their transit through the ER.

Highlights

A wide variety of proteins are known to be anchored to the extra-cytoplasmic leaflet of the plasma membrane by glycosylphosphatidylinositol (GPI) anchors and defects in GPI anchor attachment can have severe consequences for the eukaryotic cell [1]
ALS5 belongs to the agglutinin-like sequence (ALS) family of genes which code for eight adhesins in Candida albicans
In order to address the issue of whether the SS is merely a signal for GPI anchor attachment in Als5 or whether it has a say in the conformation and function of the protein, we generated the full length Als5 as well as its variant carrying the GPI-anchor signal sequence (Als5-SS) as GST-fusion products (Figure 1A) and did a comparative study of their conformation and function

Summary

Introduction

A wide variety of proteins are known to be anchored to the extra-cytoplasmic leaflet of the plasma membrane by glycosylphosphatidylinositol (GPI) anchors and defects in GPI anchor attachment can have severe consequences for the eukaryotic cell [1]. Unlike integral membrane proteins that have their transmembrane domains cotranslationally inserted into the membrane via the translocon pore, proteins meant to be GPI anchored are completely translocated into the ER lumen [3] Thereafter, these are acted upon by the GPI-transamidase and have their C-terminal GPI anchor attachment signal sequence (SS) replaced by a pre-formed GPI anchor. ALS5 belongs to the agglutinin-like sequence (ALS) family of genes which code for eight adhesins in Candida albicans These adhesins are important for establishment of commensal colonies of the organism in the host as well as in its pathogenesis and virulence under appropriate conditions [4]. Proteins destined to be Glycosylphosphatidylinositol (GPI) anchored are translocated into the ER lumen completely before the C-terminal GPI anchor attachment signal sequence (SS) is removed by the GPI-transamidase and replaced by a pre-formed GPI anchor precursor. Does the SS have a role in dictating the conformation and function of the protein as well?

Methods

Results

Conclusion