Peptide substrate conformation effect

Proline is one amino acid which would be expected to profoundly influence the reactivity of a peptide substrate. Proline restricts the possible conformations of a peptide chain and in addition is unable to act as a hydrogen bond donor. Both of these factors could affect peptide bond cleavage by hindering substrate binding or by preventing proper catalysis. Alternately, favorable interaction could take place between the enzyme and a prolyl residue, due either to a favorable hydrophobic interaction with the prolyl side chain or because proline restricts the peptide conformation to one which is favorable. That these effects are important is accentuated by the fact that at every subsite the cleavage probability of a substrate with a proline residue is significantly different from the mean (0.148). This is true for no other amino acid residue. Proline is favorable at P4 and P3 and unfavorable at all other subsites (P2 P3) ... 

With the support of quantum mechanics this proteolysis study has readily shown that fluorinated amino acid side chains are able to direct enzyme substrate interactions, which can have an influence on proteolytic stability. Depending on the absolute stereochemistry and on the position within the sequence, aTfm amino acids can considerably stabilize peptides against proteolysis. The unique electrostatic properties of carbon-bound fluorine, however, may also induce a contrary effect. The conformational restrictions of C -dialkylation seem to be partly dimin-ishable by the electrostatic consequences of fluorination. With this knowledge. 

Phosphonopeptides containing a transition state analog of the hydrolysis of the amide bond represent another attractive approach for the preparation of proteolitically stable peptides (10,30,31). In addition to increased stability, incorporation of a phosphonate moiety into the peptide sequence provides access to additional binding interactions within the transition-state conformation of the enzyme/substrate complex (13). This peptidomimetic approach is used to design very effective protease inhibitors (31-34). As in the case... 

In general, a FRET quench readout is simple. A broad range of available fluorescence donors and acceptors allows cost-efficient operations in an industrialized HTS and automated compound profiling environment. On the other hand, the readout can suffer from inner filter effects due to high absorption coefficients of the dyes and fluorescence artifacts by the tested compounds, resulting in enhanced false positive and false negative rates. Moreover, the readout is limited to substrates in which short distances between donor and acceptor dye can be realized without disturbing the interaction of enzyme and substrate. The flexibility of the peptide conformation makes the prediction of the effective distance between the dyes and consequently the prediction of the FRET effect difficult. The distance between donor and acceptor cannot be easily approximated by the mean hydrodynamic radii of the dyes. 

Anti-ribosomal antibiotics are enzyme inhibitors, and the ones that are dinically useful inhibit bacterial ribosomes far more effectively than they inhibit eukaryotic ribosomes. Many enzyme inhibitors exert their effects by binding to the active site of enzymes and thereby prevent the binding of substrates. Others block enzyme function by inhibiting conformational changes essential for enzyme activity. Anti-ribosomal antibiotics are unexceptional in this regard. They bind to the sites on both subunits to which tRNA also binds. Some block the interactions of substrates with the ribosome. Others block the tunnel and thereby prevent the nascent peptide from extending. 

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