pH-induced conformational transition in the soluble CuA domain of Paracoccus denitrificans cytochrome oxidase.


Abstract

The pH-induced conformational transition in the CuA domain of subunit II of cytochrome oxidase of Paracoccus denitrificans (PdII) has been investigated using various spectroscopic and stopped-flow kinetic methods. UV-visible absorption and circular dichroism studies showed that an increase in pH from 6 to 10 leads to a conformation change with pK(a) = 8.2 associated with the CuA site of the protein. The secondary structure of the protein was, however, shown to remain unchanged in these two conformational states. Thermal and urea-induced unfolding studies showed that the "low-pH" conformation is more stable compared to the "high-pH" conformation of the protein. Moreover, the overall stability of the protein was found to decrease on reduction of the metal centers in the low-pH form, while the oxidation state of the metal centers did not have any significant effect on the overall stability of the protein in the high-pH form. Stopped-flow pH-jump kinetic studies suggested that the conformational transition is associated with a slow deprotonation step followed by fast conformational equilibrium. The results are discussed in the light of understanding the pH-induced conformational change in the beta-barrel structure of the protein and its effect on the coordination geometry of the metal site. Study holds ProTherm entries: 11131, 11132 Extra Details: low-pH form; metal center; fast conformational equilibrium;,beta-barrel structure; coordination geometry

Submission Details

ID: xDGS6egt

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:41 p.m.

Version: 1

Publication Details
Gupta S;Warne A;Saraste M;Mazumdar S,Biochemistry (2001) pH-induced conformational transition in the soluble CuA domain of Paracoccus denitrificans cytochrome oxidase. PMID:11352755
Additional Information

Study Summary

Number of data points 6
Proteins Cytochrome c oxidase subunit 1-beta ; Cytochrome c oxidase subunit 1-beta
Unique complexes 1
Assays/Quantities/Protocols Experimental Assay: Cm buffers:borate hydrochloride: 50 mM, pH:10.0 ; Experimental Assay: m buffers:borate hydrochloride: 50 mM, pH:10.0 ; Experimental Assay: dG_H2O buffers:borate hydrochloride: 50 mM, pH:10.0 ; Experimental Assay: Cm pH:6.0, buffers:phosphate: 50 mM ; Experimental Assay: m pH:6.0, buffers:phosphate: 50 mM ; Experimental Assay: dG_H2O pH:6.0, buffers:phosphate: 50 mM
Libraries Mutations for sequence A:MADAAVHGHGDHHDTRGFFTRWFMSTNHKDIGILYLFTAGIVGLISVCFTVYMRMELQHPGVQYMCLEGARLIADASAECTPNGHLWNVMITYHGVLMMFFVVIPALFGGFGNYFMPLHIGAPDMAFPRLNNLSYWMYVCGVALGVASLLAPGGNDQMGSGVGWVLYPPLSTTEAGYSMDLAIFAVHVSGASSILGAINIITTFLNMRAPGMTLFKVPLFAWSVFITAWLILLSLPVLAGAITMLLMDRNFGTQFFDPAGGGDPVLYQHILWFFGHPEVYIIILPGFGIISHVISTFAKKPIFGYLPMVLAMAAIGILGFVVWAHHMYTAGMSLTQQAYFMLATMTIAVPTGIKVFSWIATMWGGSIEFKTPMLWAFGFLFLFTVGGVTGVVLSQAPLDRVYHDTYYVVAHFHYVMSLGAVFGIFAGVYYWIGKMSGRQYPEWAGQLHFWMMFIGSNLIFFPQHFLGRQGMPRRYIDYPVEFAYWNNISSIGAYISFASFLFFIGIVFYTLFAGKRVNVPNYWNEHADTLEWTLPSPPPEHTFETLPKREDWDRAHAH/B:MMAIATKRRGVAAVMSLGVATMTAVPALAQDVLGDLPVIGKPVNGGMNFQPASSPLAHDQQWLDHFVLYIITAVTIFVCLLLLICIVRFNRRANPVPARFTHNTPIEVIWTLVPVLILVAIGAFSLPILFRSQEMPNDPDLVIKAIGHQWYWSYEYPNDGVAFDALMLEKEALADAGYSEDEYLLATDNPVVVPVGKKVLVQVTATDVIHAWTIPAFAVKQDAVPGRIAQLWFSVDQEGVYFGQCSELCGINHAYMPIVVKAVSQEKYEAWLAGAKEEFAADASDYLPASPVKLASAE/C:EVKLQESGGDLVQPGGSLKLSCAASGFTFSSYTMSWVRQTPEKRLEWVASINNGGGRTYYPDTVKGRFTISRDNAKNTLYLQMSSLKSEDTAMYYCVRHEYYYAMDYWGQGTTVTVSSAWRHPQFGG/D:DIELTQTPVSLSASVGETVTITCRASENIYSYLAWYQQKQGKSPQFLVYNAKTLGEGVPSRFSGSGSGTQFSLKINSLLPEDFGSYYCQHHYGTPPLTFGGGTKLEIKREQKLISEEDLM

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1MQK 2003-04-01 1.28 Crystal structure of the unliganded Fv-fragment of the anti-cytochrome C oxidase antibody 7E2
3HB3 2009-06-23 2.25 High resolution crystal structure of Paracoccus denitrificans cytochrome c oxidase
3EHB 2008-09-30 2.32 A D-Pathway Mutation Decouples the Paracoccus Denitrificans Cytochrome c Oxidase by Altering the side chain orientation of a distant, conserved Glutamate
1AR1 1998-02-11 2.7 Structure at 2.7 Angstrom Resolution of the Paracoccus Denitrificans two-subunit Cytochrome C Oxidase Complexed with an Antibody Fv Fragment
1QLE 1999-12-02 3.0 CRYO-STRUCTURE OF THE PARACOCCUS DENITRIFICANS FOUR-SUBUNIT CYTOCHROME C OXIDASE IN THE COMPLETELY OXIDIZED STATE COMPLEXED WITH AN ANTIBODY FV FRAGMENT

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Cytochrome c oxidase subunit 1-beta Q00502 COX1_PARVE
90.5 Cytochrome c oxidase subunit 1-beta P08305 COX1A_PARDE
100.0 Cytochrome c oxidase subunit 1-beta P98002 COX1B_PARDE
100.0 B Cytochrome c oxidase subunit 1-beta P08306 COX2_PARDE
92.9 C Cytochrome c oxidase subunit 1-beta P18525 HVM54_MOUSE