Biologically active conformation

The second application of the CFTI protocol is the evaluation of the free energy differences between four states of the linear form of the opioid peptide DPDPE in solution. Our primary result is the determination of the free energy differences between the representative stable structures j3c and Pe and the cyclic-like conformer Cyc of linear DPDPE in aqueous solution. These free energy differences, 4.0 kcal/mol between pc and Cyc, and 6.3 kcal/mol between pE and Cyc, reflect the cost of pre-organizing the linear peptide into a conformation conducive for disulfide bond formation. Such a conformational change is a pre-requisite for the chemical reaction of S-S bond formation to proceed. The predicted low population of the cyclic-like structure, which is presumably the biologically active conformer, agrees qualitatively with observed lower potency and different receptor specificity of the linear form relative to the cyclic peptide. 

C. B. Anfinsen (Bethesda) work on ribo-nuclease, especially concerning the connection between the amino-acid sequence and the biologically active conformation. 

Teleocidines are known as tumor-promoting compounds characterized by an indolactam V core structure. Such indolactams adopt two stable conformations at room temperature. With the intention to investigate the biologically active conformer, Irie and Wender synthesized conformationally restricted analogs of indolactam V 50 (R=Me) [17]. Starting from desmethylindolactam... 

Biopharmaceuticals based on natural proteins and peptides are often called by the same name as the biologic natural material despite differences in one or more amino-acid residues. For example, insulin, which regulates blood glucose and is used clinically to treat type 1 diabetes and some cases of type 2 diabetes, has several variants that are approved for human use. Insulin contains two polypeptides, A and B chains (Figure 1.2), that are linked together by two disulfide bridges to assume a biologically active conformation. Compared with human insulin, insulin extracted from beef tissue exhibits threonine alanine and isoleucine valine substitutions at posi-... 

ANFINSEN, CHRISTIAN B. (1916-1995). An American biochemist who won the Nobel prize for chemistry- in 1972 for his work on ribonuclease, especially concerning the connection between the amino acid sequence and die biologically active conformation. He shared the Nobel prize with Stanford Moore and William H. Stein. His doctorate was granted from Harvard. 

Schematic representation of an experiment to demonstrate that the information for folding into a biologically active conformation is contained in the protein s amino acid sequence.

The sequence of amino acids in the long chain defines the primary structure of a protein. A secondary structure is determined when several residues, linked by hydrogen bonds, conform to a given combination (e.g., the a-helix, pleated sheet, and P-turns). Tertiary structure refers to the three-dimensional folded conformation of a protein. This is the biologically active conformation (crystal structure). A quaternary structure can result when two or more individual proteins assemble into two or more polypeptide chains. Conjugated proteins are complexes of proteins with other biomolecules, such as glycoproteins (sugar-proteins). 

The first consideration when setting up an AlphaScreen assay is the choice of an assay format In most cases, the decision will depend on the interaction partners under investigation and on the biological tools available for their detection. The interaction partners can be coupled to the beads directly via reductive amination of reactive aldehyde groups, similar to the immobilization on a Biacore sensor chip (see above). The usefulness of this approach is limited by the reaction conditions, which may not be appropriate for maintaining the biologically active conformation of the biomolecule. Therefore the biomolecule of interest is usually not coupled to the beads directly, but instead captured via an antibody, also preventing steric hindrance. While not strictly necessary, it is often convenient to use a biotinylated molecule which can be captured by streptavidin-coated donor beads. 

Marshall GR. Studies of the biologically active conformations of angiotensin, Vol 5. In Intra-Science Chemistry Reports. Kharasch N, ed. 1971. Gordon and Breach Science Publishers, New York, pp. 305-316. 

Finch AM, Vogen SM, Sherman S A, Kimarsky L, Taylor SM, Sanderson SD (1997) Biologically active conformer of the effector region of human C5a and modnlatory effects of N-tenninal receptor binding determinants on activity. J Med Chem 40 877-884. 

This skewed conformation also imparts further stereospecific characteristics to the hormone T3 which contains only a single outer ring iodine. Because of the restricted rotation about the two diphenyl ether bonds, the chemically equivalent 3 and 5 -iodines are conformationally distinct, giving rise to a distal (away) or proximal (near) conformation (Figure 3). To verify the importance of this conformational feature, numerous structural analogues of triiodothyronine were synthesized (4) in an effort to determine the biologically active conformer of T. ... 

---