Synthesis of functionalized de novo designed 8-16 kDa model proteins towards metal ion-binding and esterase activity.
De novo design and total chemical synthesis of proteins provides a powerful approach for biological and biophysical studies with the ability to prepare artificial proteins with tailored properties, potentially of importance for biophysical studies, material science, nanobioscience, and as molecular probes. In this paper, the previously developed concept of carbohydrates as templates is employed in the de novo design of model proteins (artificial helix bundles) termed 'carboproteins'. The 4-alpha-helix bundle is a macromolecular structure, where four amphiphilic alpha-helical peptide strands form a hydrophobic core. Here this structure is modified towards achieving metal ion-binding and catalytic activity. We report: (i) test of directional effects from different tetravalent carbohydrate templates, (ii) synthesis and evaluation of carboproteins functionalized with phenol, pyridyl or imidazolyl moieties as potential ligands for metal ion-binding as well as for catalysis. Our results include: (i) support of our previous 'controversial' finding that for some carboproteins the degree of alpha-helicity depends on the template, i.e., that there is, to some extent, a controlling effect from the template, (ii) demonstration of binding of Cu(ii) to tetra-functional carboproteins by electrospray ionization-time of flight-mass spectrometry (ESI-TOF-MS), UV-VIS absorption spectroscopy and size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS); (iii) a kinetic investigation of the esterase activity.
Pernille Tofteng
A
A
Department of Natural Sciences, Faculty of Life Sciences, University of Copenhagen, Thorvaldsensvej 40, DK-1871, Frederiksberg C, Denmark.
Hansen
Thomas H
TH
Brask
Jesper
J
Nielsen
John
J
Thulstrup
Peter W
PW
Jensen
Knud J
KJ
D000595:Amino Acid Sequence; D002241:Carbohydrates / Q000737:chemistry; D002614:Chelating Agents / Q000737:chemistry; D002942:Circular Dichroism; D020650:Combinatorial Chemistry Techniques; D003300:Copper / Q000737:chemistry; D004950:Esterases / Q000737:chemistry* / Q000378:metabolism; D006023:Glycoproteins / Q000138:chemical synthesis* / Q000737:chemistry; D013058:Mass Spectrometry; D008667:Metalloproteins / Q000138:chemical synthesis* / Q000737:chemistry; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry
D002241:Carbohydrates; D002614:Chelating Agents; D006023:Glycoproteins; D008667:Metalloproteins; D010455:Peptides; D003300:Copper; D004950:Esterases