Protein-based constraints

To determine the dosage sensitivity we calculated the packing deficiency of each gene-encoded protein based on its PDB coordinates, if available. Otherwise, packing deficiency was determined based on homology-threaded structure coordinates adopting as templates PDB-reported paralogs [19]. The input for the computation consists of the set of constraints applied to the spatial structure of the amino acid sequence to be modeled and the output is the 3D structure that best satisfies these constraints. 

For this purpose, it has been demonstrated that the high resolution solid-state NMR approach provides one with an alternative and convenient means to distinguish a variety of crystalline polymorphs and to reveal the secondary structures of biological macromolecules, because the chemical shifts of backbone carbons are displaced (up to 8 ppm) [1, 2] depending on their respective conformations. In addition, it is emphasized that this type of empirical approach can be used as a very valuable constraint to construct the three-dimensional structure of biological molecules, such as peptides and proteins, based on a set of accurately determined interatomic distances measured by a partial dipolar recoupling method, such as REDOR (rotational echo double resonance) [3-6]. 

Other constraints, more of a physical nature, which limit the choice of excipients, include solubility, stability, purity, the desired shelf life and appearance of the dried product, and special reconstitution requirements at the point of use. Thus, certain amino acids and oligopeptides, e.g. alanine and dialanine, provide good and chemically compatible stabilisers for protein-based products, but their use in practice is limited by their low solubilities. Where products are liable to exposure to high temperatures, perhaps during transport and/or storage, excipients with high Tg values are indicated. This excludes the use of amino acids and monosaccharides, even some disaccharides. Amino acids and peptides... 

As an alternative means, it is a natural consequence to expect that high-resolution sohd-state NMR could be conveniently utilized to reveal the 3D structure and dynamics of a variety of membrane proteins, because the expected NMR line widths available from sohd-state NMR are not any more influenced by motional fluctuation of proteins under consideration as a whole as encountered in solution NMR. For instance, an attempt was made to determine 3D structure of uniformly C-labeled a-spectrin SH3 domain as a globular protein, based on distance constraints estimated from proton-driven spin-diffusion (PDSD) measure-... 

Consideration of potential medical applications of protein-based polymers will always be incomplete. The source of any particular listing will be limited by confidentiality constraints on its author, by the limits of the author s imagination, and by the decision to leave out applications presumed to be out of context. At first glance, the list of medical applications included in this section 9.4 may seem long. It is, nonetheless, only a beginning. 

The amount of computation necessary to try many conformers can be greatly reduced if a portion of the structure is known. One way to determine a portion of the structure experimentally is to obtain some of the internuclear distances from two-dimensional NMR experiments, as predicted by the nuclear Over-hauser effect (NOE). Once a set of distances are determined, they can be used as constraints within a conformation search. This has been particularly effective for predicting protein structure since it is very difficult to obtain crystallographic structures of proteins. It is also possible to define distance constraints based on the average bond lengths and angles, if we assume these are fairly rigid while all conformations are accessible. 

---