title: Effects of protein engineering and rational mutagenesis on crystal lattice of single chain antibody fragments.
abstract: Protein crystallization is dependent upon, and sensitive to, the intermolecular contacts that assist in ordering proteins into a three-dimensional lattice. Here we used protein engineering and mutagenesis to affect the crystallization of single chain antibody fragments (scFvs) that recognize the EE epitope (EYMPME) with high affinity. These hypercrystallizable scFvs are under development to assist difficult proteins, such as membrane proteins, in forming crystals, by acting as crystallization chaperones. Guided by analyses of intermolecular crystal lattice contacts, two second-generation anti-EE scFvs were produced, which bind to proteins with installed EE tags. Surprisingly, although noncomplementarity determining region (CDR) lattice residues from the parent scFv framework remained unchanged through the processes of protein engineering and rational design, crystal lattices of the derivative scFvs differ. Comparison of energy calculations and the experimentally-determined lattice interactions for this basis set provides insight into the complexity of the forces driving crystal lattice choice and demonstrates the availability of multiple well-ordered surface features in our scFvs capable of forming versatile crystal contacts.
authors.affiliation: School of Chemistry & Biochemistry, Georgia Institute of Technology, Atlanta, Georgia, 30332-0400.
authors.forename: Sibel
authors.initials: S
authors.lastname: Kalyoncu
authors.forename: Jeongmin
authors.initials: J
authors.lastname: Hyun
authors.forename: Jennifer C
authors.initials: JC
authors.lastname: Pai
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Johnson
authors.forename: Kevin
authors.initials: K
authors.lastname: Entzminger
authors.forename: Avni
authors.initials: A
authors.lastname: Jain
authors.forename: David P
authors.initials: DP
authors.lastname: Heaner
authors.forename: Ivan A
authors.initials: IA
authors.lastname: Morales
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Truskett
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Maynard
authors.forename: Raquel L
authors.initials: RL
authors.lastname: Lieberman
mesh_terms: D000595:Amino Acid Sequence; D019295:Computational Biology; D003460:Crystallization; D018360:Crystallography, X-Ray; D000939:Epitopes / Q000235:genetics*; D008958:Models, Molecular; D018832:Molecular Chaperones; D016296:Mutagenesis; D011485:Protein Binding; D015202:Protein Engineering; D060066:Protein Interaction Maps; D011506:Proteins / Q000235:genetics*; D057127:Single-Chain Antibodies / Q000235:genetics*
chemical_list: D000939:Epitopes; D018832:Molecular Chaperones; D011506:Proteins; D057127:Single-Chain Antibodies
keywords: crystal packing; intermolecular contacts; protein crystallography; protein interaction; scFv affinity; short peptide epitope