title: Heteromeric assembled polypeptidic artificial hydrolases with a six-helical bundle scaffold.
abstract: Enzyme efficiency results from the cooperation of functional groups in the catalytic site. In order to mimic a natural enzyme, a definite 3D scaffold must be carefully designed so that the functional groups can work cooperatively. During the HIV-1 fusion process, the gp41 N- and C-terminal heptad repeat regions form a coiled-coil six-helical bundle (6HB) that brings the viral and target cell membranes into close proximity for fusion. We used 6HB as the molecular model for a novel scaffold for the design of an artificial enzyme, in which the modified C34 and N36 peptides formed a unique 6HB structure through specific molecular recognition, and the position and orientation of the side-chain groups on this scaffold were predictable. The histidine modified 6HB C34(H13/20)/N36(H15/22) showed enzyme-like hydrolytic activity towards p-nitrophenyl acetate (PNPA; k(cat)/K(M) =3.66 M(-1) s(-1)) through the cooperation of several inter- or intrahelical imidazole groups. Since the catalytic activity of 6HB depends on the C- and N-peptide assembly, either HIV fusion inhibitors that can compete with the formation of catalytic 6HB or denaturants that can destroy the ordered structure were able to modulate its activity. Further engineering of the solvent-exposing face with Glu(-)-Lys(+) salt bridges enhanced the helicity and the stability of 6HB. As a result, the population and stability of cooperative catalytic units increased. In addition, the Glu(-)-Lys(+) -stabilized 6HB SC35(H13/20)/N36(H15/22) had increased catalytic efficiency (k(cat)/K(M) =6.30 M(-1) s(-1)). A unique 6HB system was specifically assembled and provided a scaffold sufficiently stable to mimic the function of enzymes or other biomolecules.
authors.affiliation: Key Laboratory of Functional Polymer Materials of MOE, Institute of Polymer Chemistry, Nankai University, Tianjin 300071, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Bai
authors.forename: Yanbo
authors.initials: Y
authors.lastname: Ling
authors.forename: Weiguo
authors.initials: W
authors.lastname: Shi
authors.forename: Lifeng
authors.initials: L
authors.lastname: Cai
authors.forename: Qiyan
authors.initials: Q
authors.lastname: Jia
authors.forename: Shibo
authors.initials: S
authors.lastname: Jiang
authors.forename: Keliang
authors.initials: K
authors.lastname: Liu
mesh_terms: D000595:Amino Acid Sequence; D055162:Biocatalysis; D040761:Biomimetic Materials / Q000737:chemistry / Q000378:metabolism*; D002942:Circular Dichroism; D015700:HIV Envelope Protein gp41 / Q000737:chemistry / Q000378:metabolism; D023581:HIV Fusion Inhibitors / Q000737:chemistry / Q000494:pharmacology; D015497:HIV-1 / Q000378:metabolism; D006801:Humans; D006867:Hydrolases / Q000737:chemistry / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D009596:Nitrophenols / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D015202:Protein Engineering; D017433:Protein Structure, Secondary
chemical_list: D015700:HIV Envelope Protein gp41; D023581:HIV Fusion Inhibitors; D009596:Nitrophenols; D010446:Peptide Fragments; C407149:peptide C34; C407148:peptide N36; C008642:4-nitrophenyl acetate; D006867:Hydrolases