Solid peptides

Modern combinatorial chemistry involves a number of steps that begin with the creation of a library of molecules that are closely related in structure. The library can be created in two ways (a) parallel synthesis, which is simultaneous synthesis of numerous products in separate discrete reaction vessels (b) combinatorial synthesis, of numerous reactions within one single reaction vessel followed by separations. The initial successes in parallel synthesis have been in solid peptide synthesis of proteins, which was based on Merrifield s solid-phase peptide synthesis. 

M. Erbeldinger, X. Ni, and P. J. Halling, Effect of water and enzyme concentration on thermolysin-catalyzed solid-to-solid peptide synthesis, Biotechnol. Bioeng. 1998, 59, 68-72. 

Kuroki, S., Takahashi, A., Ando, I., Shoji, A., and Ozaki, T., Hydrogen-bonding structural study of solid peptides and polypeptides containing a glycine residue by NMR spectroscopy, J. Mol. Struct. 323, 197-208 (1995). 

Kuroki, S., Ando, S., and Ando, I., An MO study of nuclear quadrupolar coupling constant and nuclear. shielding of the carbonyl oxygen in solid peptides with hydrogen bonds, Chem. Phys. 195, 107-116(1995). 

Dialyze the carrier protein against 10 mM potassium phosphate buffer, pH 7.2. At ust the carrier concentration to about 20 mg/mL. Weigh out the solid peptide and dissolve it in the same buffer at 5 mg/mL. Thiol-based scavengers, used to preserve free -SH groups in the peptide, are often present in peptide preparations and will need to be removed from the peptide solution by desalting on a Bio-Gel PIO column in 10 mM potassium phosphate buffer, pH 7.2. A brief nasal inspection is often sufficient to determine their presence. On the other hand, stock solutions of peptide (e.g., in PBS) may become oxidized if stored for extended periods, and so are best pretreated with 100 mMDTT (30 min at room temperature is adequate) to reduce the dissolved peptide before use. As the DTT will interfere with the conjugation, the reduced peptide should then be desalted on a PIO column as above. 

Table 6.5.1. Determined NMR parameters of solid peptides containing Gly residue...

Chaiwut P, Kanasawud P, Hailing PJ (2007) Solid-to-solid peptide synthesis by glycyl endopepti-dase. Enzyme Microb Technol 40(4) 954-960... 

Spectra of solid peptides Ccin be used to describe conformational features of the peptides. Of particular utility are the Pro and Cy chemical shifts, which can describe ring geometry or occurrence of cis or trans X-Pro bond conformation. 

To improve the access to the functional groups, Lee and coworkers developed a process of placing most of the functional groups on the outer surface of the polystyrene beads [7]. This was done by surface aminomethylation of preformed cross-Unked commercially available polystyrene beads. Such bead usually range in size from 100 to 400 mesh (in.) in diameter and can be functionalized by surface reactions or by surface grafts. This yielded materials with a lower number of functional groups, but with the majority of them being accessible. Lee and coworkers then utilized the beads in a solid peptide synthesis [7]. 

Most studies of internal dynamics in solid peptides and proteins, exploited the and N Tj relaxation times measurements thus, the time scale is limited to the range from picoseconds to microseconds. In order to extend the range of observable dynamic processes, one has to employ more sophisticated NMR approaches. 

The ultraslow motions can be observed by exchange NMR experiments. There are a number of techniques, performed in ID and 2D modes which are based on the manipulation of spinning sidebands under MAS. The first of them, applied to study of solid peptides was ODESSA (one-dimensional exchange spectroscopy by sideband-alternation) [76] and next the time-reversed ODESSA [77] (tr-ODESSA). The interesting modification, which represents this group of sequences is CODEX (centreband-only detection of exchange) introduced by Schmidt-Rohr and coworkers [78]. CODEX can be executed at any MAS speed, while ODESSA and tr-ODESSA... 

First-principles calculations of NMR parameters for solid peptides have a long-standing history. Number of spectacular results was pubhshed during last 20 years. Review articles of Sitkoff et al. and Ando and coworkers summarize the apphcabihty and power of GIAO approach for study of different peptides at that time [183,184]. Today, the modem methodologies are rather oriented on appHcations of GIPAW method. As shown by Zeng... 

Yan, C., Mackay, M.E., Czymmek, K., et al. Injectable solid peptide hydrogel as a cell carrier. effects of shear flow on hydrogels and cell payload. Langmuir 28, 6076-6087 (2012). doi 10.1021/la2041746... 

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