Combined solid‐phase and solution approach for the synthesis of large peptides or proteins
In the synthesis of large peptides or proteins, highly homogeneous segments are indispensable for a convergent strategy either on a solid‐phase resin or in solution. Employing Boc/Bzl chemistry to prepare fully protected segments with a free α‐carboxyl group from the solid support, base‐labile linkers are profitable for practical peptide synthesis since they require no special equipment. For this purpose, an N‐[9‐(hydroxymethyl)‐2‐fluorenyl]succinamic acid (HMFS) linker was adopted. Consequently, there must be high compatibility between the protecting groups of the segment and the anchoring group which is cleavable by treatment with morpholine or piperidine in DMF. Instead of using the 2‐bromobenzyloxycarbonyl (BrZ) group for the Tyr residue and the formyl (For) group for the Trp residue, both of which are the most susceptible protecting groups under these base‐catalysed conditions, the base‐resistant 3‐pentyl (Pen) and cyclohexyloxycarbonyl (Hoc) groups were introduced to the respective side‐chain functional groups. By applying the present strategy, the authors were able to rapidly synthesize homogeneous protected segments for use in the subsequent segment coupling in solution. In the present paper, the utility of the combined solid‐phase and solution approach is demonstrated by synthesizing muscarinic toxin 1 (MTX1) which binds to the muscarinic acetylcholine receptors. Copyright © 2000 European Peptide Society and John Wiley & Sons, Ltd.
3-Pentyl (Pen) group as a new base-resistant side chain protecting group for tyrosine
Abstract The 3-pentyl (Pen) group is a new base-resistant protecting group for tyrosine. It is sufficiently stable in 50% TFA/DCM and completely stable towards 20% piperidine/DMF but is readily cleavable by the standard HF procedure without the formation of any significant amount of alkyltyrosine rearrangement product.
Two new organic reference materials for delta13C and delta15N measurements and a new value for the delta13C of NBS 22 oil.
Analytical grade L-glutamic acid is chemically stable and has a C/N mole ratio of 5, which is close to that of many of natural biological materials, such as blood and animal tissue. Two L-glutamic acid reference materials with substantially different 13C and 15N abundances have been prepared for use as organic reference materials for C and N isotopic measurements. USGS40 is analytical grade L-glutamic acid and has a delta13C value of -26.24 per thousand relative to VPDB and a delta15N value of -4.52 per thousand relative to N2 in air. USGS41 was prepared by dissolving analytical grade L-glutamic acid with L-glutamic acid enriched in 13C and 15N. USGS41 has a delta13C value of +37.76 per thousand and a delta15N value of +47.57 per thousand. The delta13C and delta15N values of both materials were measured against the international reference materials NBS 19 calcium carbonate (delta13C=+1.95 per thousand ), L-SVEC lithium carbonate (delta13C=-46.48 per thousand ), IAEA-N-1 ammonium sulfate (delta15N=0.43 per thousand ), and USGS32 potassium nitrate (delta15N=180 per thousand ) by on-line combustion continuous-flow and off-line dual-inlet isotope-ratio mass spectrometry. Both USGS40 and USGS41 are isotopically homogeneous; reproducibility of delta13C is better than 0.13 per thousand, and that of delta15N is better than 0.13 per thousand in 100-microg amounts. These two isotopic reference materials can be used for (i) calibrating local laboratory reference materials, and (ii) quantifying drift with time, mass-dependent fractionations, and isotope-ratio-scale contraction in the isotopic analysis of various biological materials. Isotopic results presented in this paper yield a delta13C value for NBS 22 oil of -29.91 per thousand, in contrast to the commonly accepted value of -29.78 per thousand for which off-line blank corrections probably have not been quantified satisfactorily.