Search results for "Cofactor binding"
showing 4 items of 4 documents
SDS-facilitated in vitro formation of a transmembrane B-type cytochrome is mediated by changes in local pH.
2011
Abstract The folding and stabilization of α-helical transmembrane proteins are still not well understood. Following cofactor binding to a membrane protein provides a convenient method to monitor the formation of appropriate native structures. We have analyzed the assembly and stability of the transmembrane cytochrome b 559 ′, which can be efficiently assembled in vitro from a heme-binding PsbF homo-dimer by combining free heme with the apo-cytochrome b 559 ′. Unfolding of the protein dissolved in the mild detergent dodecyl maltoside may be induced by addition of SDS, which at high concentrations leads to dimer dissociation. Surprisingly, absorption spectroscopy reveals that heme binding and…
A Ser residue influences the structure and stability of a Pro-kinked transmembrane helix dimer
2012
AbstractWhen localized adjacent to a Pro-kink, Thr and Ser residues can form hydrogen bonds between their polar hydroxyl group and a backbone carbonyl oxygen and thereby modulate the actual bending angle of a distorted transmembrane α-helix. We have used the homo-dimeric transmembrane cytochrome b559′ to analyze the potential role of a highly conserved Ser residue for assembly and stabilization of transmembrane proteins. Mutation of the conserved Ser residue to Ala resulted in altered heme binding properties and in increased stability of the holo-protein, most likely by tolerating subtle structural rearrangements upon heme binding. The results suggest a crucial impact of an intrahelical Ser…
Crystal Structure of Perakine Reductase, Founding Member of a Novel Aldo-Keto Reductase (AKR) Subfamily That Undergoes Unique Conformational Changes …
2012
Perakine reductase (PR) catalyzes the NADPH-dependent reduction of the aldehyde perakine to yield the alcohol raucaffrinoline in the biosynthetic pathway of ajmaline in Rauvolfia, a key step in indole alkaloid biosynthesis. Sequence alignment shows that PR is the founder of the new AKR13D subfamily and is designated AKR13D1. The x-ray structure of methylated His(6)-PR was solved to 2.31 Å. However, the active site of PR was blocked by the connected parts of the neighbor symmetric molecule in the crystal. To break the interactions and obtain the enzyme-ligand complexes, the A213W mutant was generated. The atomic structure of His(6)-PR-A213W complex with NADPH was determined at 1.77 Å. Overal…
Structural dissection of the multidomain kininogens. Fine mapping of the target epitopes of antibodies interfering with their functional properties.
1993
Kininogens, the large precursor molecules of the vasoactive kinin peptides, are prototypic multidomain proteins serving numerous functions. To investigate their structure-function relationships, we have raised a panel of monoclonal antibodies against human H-kininogen and L-kininogen and fragments thereof and characterized them with respect to their target epitopes. Of 35 antibodies, 12 were directed to the amino-terminal domains (D1 to D3) of cystatin-like structure, 3 recognized domain D4 bearing the kinin segment, 17 bound to the carboxyl-terminal domains of H-kininogen (D5H and D6H), and 3 bound to the carboxyl-terminal domain D5L of L-kininogen. At least 14 distinct epitopes spread ove…