A case of spurious product formation during attempted resynthesis of proteins by reverse proteolysis. Some batches of ‘pure’ glycerol contain cross-linking agents

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Abstract
In cases is which enzyme-catalyzed synthesis of a peptide bond is being used to re-form a protein from 2 large peptide fragments, the organic co-solvent chosen has so far been glycerol for most solvents in use in small-molecule systems are potent protein denaturants. Impurities contaminating certain batches of glycerol are effective in cross-linking the complexes formed by these peptide fragments, mimicking the enzyme-catalyzed process. In one such case, the reported re-formation of cytochrome c from a 2-fragment complex system, cytochrome c-T, the extent and rate of conjugate formation duplicates that reported for enzymic resynthesis. No difference between mixtures containing or lacking enzyme was observed. The danger of confusion possible to those engaged in studies of resynthesis was discussed, and a simple control of purchased glycerol was suggested to avoid it. Similar caution to those (X-ray crystallographers and others) who seek to stabilize protein solutions by adding large quantities of glycerol is recommended.