  Accurate treatment of amide resonance is important in electronic structure
calculation of protein, for Resonance-Assisted Hydrogen Bonding in the hydrogen
bonded chains of backbone amides of protein secondary structures such as beta
sheets and alpha helices is determined by amide resonance. Variation in amide
resonance is the means by which the hydrogen bonding in these chains is
cooperative.
  Amide carbonyl orbitals are revealed by Natural Bond Orbital, NBO, analysis
to substantially maintain sigma/pi separation in the presence of torsional
hyperconjugative interactions with wavefunction methods but not with
established Density Functional Theory, DFT, methods. This DFT error is most
pronounced with small basis sets such as are used with DFT for proteins to
reduce the basis function count. This error disturbs calculation of a range of
amide donor-acceptor and steric interactions.
  This finding has important implications for the selection of electronic
structure methods and basis sets for protein calculations. For example, great
caution is needed in interpreting the results of applying established DFT
methods to proteins containing any beta sheets. We recommend that every protein
DFT calculation be accompanied by NBO assessment of maintenance of amide
carbonyl sigma/pi separation and absence of carbonyl bond bending. Further, we
propose that these metrics be standard benchmarks of electronic structure
methods and basis sets.
