TY - JOUR
T1 - Protein Conformational Change Delayed by Steric Hindrance from an N-Linked Glycan
AU - Bager, René
AU - Johansen, Jesper Sanderhoff
AU - Jensen, Jan K.
AU - Stensballe, Allan
AU - Jendroszek, Agnieszka
AU - Buxbom, Linette
AU - Sørensen, Hans Peter
AU - Andreasen, Kristian Peter
N1 - Copyright © 2013. Published by Elsevier Ltd.
PY - 2013/5/20
Y1 - 2013/5/20
N2 - Very few studies have attributed a direct, active, functional role to N-linked glycans. We describe here an N-linked glycan with a unique role for maintaining the active conformation of a protein of the serpin family. The distinguishing feature of serpins is the "stressed-to-relaxed" transition, in which the reactive centre loop (RCL) inserts as a β-strand into the central β-sheet A. This transition forms the basis for the conversion of serpins to the inactive latent state. We demonstrate that plasminogen activator inhibitor-1 (PAI-1) from zebrafish converts to the latent state about 5-fold slower than human PAI-1. In contrast to human PAI-1, fish PAI-1 carries a single N-linked glycan at Asn185 in the gate region through which the RCL passes during latency transition. While the latency transition of human PAI-1 is unaffected by deglycosylation, deglycosylated zebrafish PAI-1 goes latent about 50-fold faster than the glycosylated zebrafish PAI-1 and about 25-fold faster than non-glycosylated human PAI-1. X-ray crystal structure analysis of glycosylated fish PAI-1 confirmed the presence of an N-linked glycan in the gate region and a lack of glycan-induced structural changes. Thus, latency transition of zebrafish PAI-1 is delayed by steric hindrance from the glycan in the gate region. Our findings reveal a previously unknown mechanism for inhibition of protein conformational changes by steric hindrance from N-linked glycans.
AB - Very few studies have attributed a direct, active, functional role to N-linked glycans. We describe here an N-linked glycan with a unique role for maintaining the active conformation of a protein of the serpin family. The distinguishing feature of serpins is the "stressed-to-relaxed" transition, in which the reactive centre loop (RCL) inserts as a β-strand into the central β-sheet A. This transition forms the basis for the conversion of serpins to the inactive latent state. We demonstrate that plasminogen activator inhibitor-1 (PAI-1) from zebrafish converts to the latent state about 5-fold slower than human PAI-1. In contrast to human PAI-1, fish PAI-1 carries a single N-linked glycan at Asn185 in the gate region through which the RCL passes during latency transition. While the latency transition of human PAI-1 is unaffected by deglycosylation, deglycosylated zebrafish PAI-1 goes latent about 50-fold faster than the glycosylated zebrafish PAI-1 and about 25-fold faster than non-glycosylated human PAI-1. X-ray crystal structure analysis of glycosylated fish PAI-1 confirmed the presence of an N-linked glycan in the gate region and a lack of glycan-induced structural changes. Thus, latency transition of zebrafish PAI-1 is delayed by steric hindrance from the glycan in the gate region. Our findings reveal a previously unknown mechanism for inhibition of protein conformational changes by steric hindrance from N-linked glycans.
U2 - 10.1016/j.jmb.2013.05.007
DO - 10.1016/j.jmb.2013.05.007
M3 - Journal article
C2 - 23702291
SN - 0022-2836
VL - 425
SP - 2867
EP - 2877
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 16
ER -