Cargando…
Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin
Bovine α-lactalbumin (αLA) has four disulfide (SS) bonds in the native form (N). On the oxidative folding pathways of this protein, two specific SS folding intermediates, i.e., (61–77, 73–91) and des[6–120], which have two and three native SS bonds, respectively, accumulate predominantly in the pres...
Autores principales: | , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2017
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5618645/ https://www.ncbi.nlm.nih.gov/pubmed/28926961 http://dx.doi.org/10.3390/ijms18091996 |
_version_ | 1783267235598434304 |
---|---|
author | Shinozaki, Reina Iwaoka, Michio |
author_facet | Shinozaki, Reina Iwaoka, Michio |
author_sort | Shinozaki, Reina |
collection | PubMed |
description | Bovine α-lactalbumin (αLA) has four disulfide (SS) bonds in the native form (N). On the oxidative folding pathways of this protein, two specific SS folding intermediates, i.e., (61–77, 73–91) and des[6–120], which have two and three native SS bonds, respectively, accumulate predominantly in the presence of Ca(2+). In this study, we reinvestigated the pathways using a water-soluble cyclic selenoxide reagent, trans-3,4-dihydroxyselenolane oxide (DHS(ox)), as a strong and quantitative oxidant to oxidize the fully reduced form (R). In the presence of ethylenediaminetetraacetic acid (EDTA) (under a metal-free condition), SS formation randomly proceeded, and N did not regenerate. On the other hand, two specific SS intermediates transiently generated in the presence of Ca(2+). These intermediates could be assigned to (61–77, 73–91) and des[6–120] having two common SS bonds, i.e., Cys61-Cys77 and Cys73-Cys91, near the calcium binding pocket of the β-sheet domain. Much faster folding to N was observed in the presence of Mn(2+), whereas Na(+), K(+), Mg(2+), and Zn(2+) did not affect the pathways. The two key intermediates were susceptible to temperature and a denaturant. The oxidative folding pathways revealed were significantly different from those of hen egg white lysozyme, which has the same SS-bonding pattern as αLA, suggesting that the folding pathways of SS-containing proteins can alter depending on the amino acid sequence and other factors, even when the SS-bond topologies are similar to each other. |
format | Online Article Text |
id | pubmed-5618645 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-56186452017-09-30 Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin Shinozaki, Reina Iwaoka, Michio Int J Mol Sci Article Bovine α-lactalbumin (αLA) has four disulfide (SS) bonds in the native form (N). On the oxidative folding pathways of this protein, two specific SS folding intermediates, i.e., (61–77, 73–91) and des[6–120], which have two and three native SS bonds, respectively, accumulate predominantly in the presence of Ca(2+). In this study, we reinvestigated the pathways using a water-soluble cyclic selenoxide reagent, trans-3,4-dihydroxyselenolane oxide (DHS(ox)), as a strong and quantitative oxidant to oxidize the fully reduced form (R). In the presence of ethylenediaminetetraacetic acid (EDTA) (under a metal-free condition), SS formation randomly proceeded, and N did not regenerate. On the other hand, two specific SS intermediates transiently generated in the presence of Ca(2+). These intermediates could be assigned to (61–77, 73–91) and des[6–120] having two common SS bonds, i.e., Cys61-Cys77 and Cys73-Cys91, near the calcium binding pocket of the β-sheet domain. Much faster folding to N was observed in the presence of Mn(2+), whereas Na(+), K(+), Mg(2+), and Zn(2+) did not affect the pathways. The two key intermediates were susceptible to temperature and a denaturant. The oxidative folding pathways revealed were significantly different from those of hen egg white lysozyme, which has the same SS-bonding pattern as αLA, suggesting that the folding pathways of SS-containing proteins can alter depending on the amino acid sequence and other factors, even when the SS-bond topologies are similar to each other. MDPI 2017-09-16 /pmc/articles/PMC5618645/ /pubmed/28926961 http://dx.doi.org/10.3390/ijms18091996 Text en © 2017 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Shinozaki, Reina Iwaoka, Michio Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin |
title | Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin |
title_full | Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin |
title_fullStr | Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin |
title_full_unstemmed | Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin |
title_short | Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin |
title_sort | effects of metal ions, temperature, and a denaturant on the oxidative folding pathways of bovine α-lactalbumin |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5618645/ https://www.ncbi.nlm.nih.gov/pubmed/28926961 http://dx.doi.org/10.3390/ijms18091996 |
work_keys_str_mv | AT shinozakireina effectsofmetalionstemperatureandadenaturantontheoxidativefoldingpathwaysofbovinealactalbumin AT iwaokamichio effectsofmetalionstemperatureandadenaturantontheoxidativefoldingpathwaysofbovinealactalbumin |