Water-Soluble Chlorophyll Protein (WSCP) Stably ... - ACS Publications

Mar 2, 2017 - via absorption spectroscopy using an extinction coefficient. (ε280) of 25440 ...... acknowledges the Ing. Aldo Gini foundation for supp...
0 downloads 0 Views 1MB Size
Subscriber access provided by University of Newcastle, Australia

Article

Water-Soluble Chlorophyll Protein (WSCP) Stably Binds 2 or 4 Chlorophylls Daniel M. Palm, Alessandro Agostini, Stefan Tenzer, Barbara M. Gloeckle, Mara Werwie, Donatella Carbonera, and Harald Paulsen Biochemistry, Just Accepted Manuscript • DOI: 10.1021/acs.biochem.7b00075 • Publication Date (Web): 02 Mar 2017 Downloaded from http://pubs.acs.org on March 6, 2017

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

Biochemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 38

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Biochemistry

Water-Soluble Chlorophyll Protein (WSCP) Stably Binds 2 or 4 Chlorophylls Daniel M. Palm§‡, Alessandro Agostini§†‡, Stefan Tenzer∥, Barbara M. Gloeckle§, Mara Werwie§, Donatella Carbonera†, and Harald Paulsen§* §

Institute of General Botany, Johannes-Gutenberg University Mainz, Johannes-von-Müller-Weg 6,

55128 Mainz, Germany, †Department of Chemical Sciences, University of Padova, Via Marzolo 1, 35131 Padova, Italy, ∥ Institute for Immunology, University Medical Center Mainz, Langenbeckstr. 1, 55131 Mainz, Germany. KEYWORDS: Chlorophyll; Heat stability; Photostability; pH stability; Substoichiometric complex; Water-soluble chlorophyll protein (WSCP);

ABSTRACT: Water-soluble chlorophyll proteins (WSCP) of class-IIa from Brassicaceae form tetrameric complexes containing one chlorophyll (Chl) per apoprotein but no carotenoids. The complexes are remarkably stable towards dissociation and protein denaturation even at 100 °C and extreme pH values, and the Chls are partially protected against photooxidation. There are several hypotheses for the biological role of WSCPs, one of them proposing them to function as a scavenger of Chls set free upon plant senescence or pathogen attack. The biochemical WSCP properties described in this paper are consistent with the protein acting as an efficient and flexible Chl scavenger. Under limiting Chl concentrations, the recombinant WSCP apoprotein binds substoichiometric amounts of Chl 2

ACS Paragon Plus Environment

Biochemistry

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 2 of 38

(2 Chls/tetramer) to form complexes that are as stable towards thermal dissociation, denaturation, and photodamage as the fully pigmented ones. If more Chl is added, these 2-Chl complexes can bind another 2 Chls to reach the fully pigmented state. The protection of WSCP Chls against photodamage has been attributed to the apoprotein serving as a diffusion barrier for oxygen, preventing its access to tripletexcited Chls and thus, the formation of singlet oxygen. By contrast, the sequential binding of Chls by WSCP suggests a partially open or at least flexible structure, raising the question of how WSCP photoprotects its Chls without the help of carotenoids.

3

ACS Paragon Plus Environment

Page 3 of 38

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Biochemistry

Water-soluble chlorophyll protein (WSCP) is an unusual protein in several respects. Whereas all other chlorophyll (Chl) proteins in higher plants are membrane-bound, WSCP is not. WSCP contains no carotenoids and even so, is able to protect its Chl to some extent against photooxidation.1 And finally, WSCP is extremely stable towards thermal dissociation and denaturation.1–5 This latter property is particularly intriguing as the biological function of WSCP is unclear. In this paper, we show that the stability of WSCP extends to a version with a substoichiometric number of Chls bound. All WSCPs are tetrameric with four subunits of roughly 20 kDa.6–8 Two classes of WSCPs have been described. Class-I WSCPs isolated from Amaranthaceae (including Chenopodiaceae) and Polygonaceae have in common that they shift their absorption spectra upon illumination.9 Class-II WSCPs, dealt with in this paper, show no such photoconvertibility10 and have been found so far only in Brassicaceae. The class-II WSCPs can be further subdivided into class-IIa (including Arabidopsis, Brassica, Raphanus) with a Chl a/b ratio >6, and class-IIb (so far only found in Lepidium virginicum) with Chl a/b