Zinc ion-induced domain organization in metallo-beta-lactamases: a flexible "zinc arm" for rapid metal ion transfer?
The reversible unfolding of metallo-beta-lactamase from Chryseobacterium meningosepticum (BlaB) by guanidinium hydrochloride is best described by a three-state model including folded, intermediate, and unfolded states. The transformation of the folded apoenzyme into the intermediate state requires only very low denaturant concentrations, in contrast to the Zn2-enzyme. Similarly, circular dichroism spectra of both BlaB and metallo-beta-lactamase from Bacillus cereus 569/H/9 (BcII) display distinct differences between metal-free and Zn2-enzymes, indicating that the zinc ions affect the folding of the proteins, giving a larger alpha-helix content. To identify the regions of the protein involved in this zinc ion-induced change, a hydrogen deuterium exchange study with matrix-assisted laser desorption ionization tandem time of flight mass spectrometry on metal-free and Zn1- and Zn2-BcII was carried out. The region spanning the metal binding metallo-beta-lactamases (MBL) superfamily consensus sequence His-X-His-X-Asp motif and the loop connecting the N- and C-terminal domains of the protein undergoes a zinc ion-dependent structural change between intrinsically disordered and ordered states. The inherent flexibility even appears to allow for the formation of metal ion-bridged protein-protein complexes which may account for both electrospray ionization-mass spectroscopy results obtained upon variation of the zinc/protein ratio and stoichiometry-dependent variations of 199mHg-perturbed angular correlation of gamma-rays spectroscopic data. We suggest that this flexible "zinc arm" motif, present in all the MBL subclasses, is disordered in metal-free MBLs and may be involved in metal ion acquisition from zinc-carrying molecules different from MBL in an "activation on demand" regulation of enzyme activity.
Selevsek
Nathalie
N
From the Departments of Biochemical Engineering, 66041 Saarbrücken, Germany.
Rival
Sandrine
S
Biochemistry, Saarland University, 66041 Saarbrücken, Germany.
Tholey
Andreas
A
From the Departments of Biochemical Engineering, 66041 Saarbrücken, Germany; Institute for Experimental Medicine-Systemic Proteome Research and Bioanalytics, Christian-Albrechts Universität, 24105 Kiel, Germany.
Heinzle
Elmar
E
From the Departments of Biochemical Engineering, 66041 Saarbrücken, Germany.
Heinz
Uwe
U
Department of Basic Sciences and Environment, Faculty of Life Sciences, University of Copenhagen, DK-1871 Frederiksberg C, Denmark.
Hemmingsen
Lars
L
Department of Basic Sciences and Environment, Faculty of Life Sciences, University of Copenhagen, DK-1871 Frederiksberg C, Denmark.
Adolph
Hans W
HW
Department of Basic Sciences and Environment, Faculty of Life Sciences, University of Copenhagen, DK-1871 Frederiksberg C, Denmark. Electronic address: hwa@life.ku.dk.
D001409:Bacillus cereus / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D045247:Chryseobacterium / Q000201:enzymology*; D002942:Circular Dichroism; D008956:Models, Chemical; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity; D015032:Zinc / Q000737:chemistry* / Q000378:metabolism; D001618:beta-Lactamases / Q000737:chemistry* / Q000378:metabolism
D001426:Bacterial Proteins; C455979:beta-lactamase BcII; C410343:BLAB3 protein, Flavobacterium meningosepticum; D001618:beta-Lactamases; D015032:Zinc