Chemical Solid Phase Synthesis Raw Materials Amino Acids with Protective Groups

Classification of twenty common amino acids

The 20 kinds of amino acids can be divided into 4 groups according to their physical and chemical properties:

① Non accessible and hydrophobic amino acids: glycine (G), alanine (A), valine (V), leucine (L), toliunine (M), isoleucine (I), phenylalanine (F) and proline (P). Supplement: Methionine is also called methionine (M).

② Suitable and neutral amino acids: tryptophan (W), serine (S), tyrosine (Y), cysteine (C), asparagine (N), glutamine (Q) and threonine (T).

③ Acidic amino acids: aspartic acid (D) and glutamic acid (E).

④ Basic amino acids: lysine (K), arginine (R) and histidine (H).

(Hint: multiple hydrophobic amino acids appear continuously in the polypeptide sequence, which is a difficult point in solid phase synthesis. Because hydrophobic amino acids will allow the carrier resin of solid phase synthesis to condense, so that the amino acid sites are closed. The connecting sites are closed, and subsequent amino acids can not be connected normally. As polypeptide sequences of the same length, the synthesis of more hydrophobic amino acids is more difficult.)

Differences between synthetic amino acids and natural amino acids

Synthetic peptides are different from the synthesis of natural proteins. Artificial solid phase synthesis is generally connected by adding amino acids in sequence from the C end to the N end. In order to reduce the generation of side reactions in synthesis, the amino acids used are all protected groups, which protect the active groups that are not used temporarily in synthesis. And the common 20 kinds of natural amino acids are L-amino acids. Besides L-amino acids, there are also D-amino acids for artificial synthesis. This allows artificial synthesis of polypeptide sequences that do not exist in the natural world.

Amino acid protective group for artificial synthesis.

Common main chain protection bases: Fmoc, Boc.

Common side chain protection bases: Boc, Trt, OtBu, tBu, Pbf.

Examples of amino acids used for synthesis:

Fmoc-Ala-OH; (Fmoc refers to protecting the amino group of the main chain)

Fmoc-Cys（Trt）-OH; (Parentheses indicate the temporary protective group of the active group protecting the side chain)

Fmoc-Lys（Boc）-OH;

Fmoc-Ser(tBu)-OH;

Fmoc-Asp（otBu）-OH

Fmoc-Arg(Pbf)-OH.

Boc-Cys（Trt）-OH

Fmoc Ala OH structural formula

How to use amino acids with protective groups to synthesize polypeptide sequences

Take the Fmoc protection strategy method as an example. Synthesis consists of the following steps:

① Hanging resin: select and weigh the corresponding amount of resin according to the order, and put it into the reactor for swelling and standby.

② Deprotection: The column and monomer protected by Fmoc must use 20% piperidine to remove the protective group of amino group.

③ Detection: detect whether the protective group has been removed, i.e. whether the amino acid has been exposed. (Ontolex Biology often uses ninhydrin as the test reagent. If the result is positive, it will be judged as amino fruit juice)

④ Activation and cross-linking: The carboxyl group of the next amino acid is activated by an activator. The activated monomer is cross-linked with free amino to form peptide bond. We need to note that the amount of amino acid input is not arbitrary. To ensure the accuracy of peptide chain synthesis; Amino acids, condensation agents and bases have a certain proportion. A reference scale is provided here. (Scale (mol): amino acid: condensation agent: alkali=1:3:3:6)

⑤ Test: check whether the required amino acid has been connected. The result is negative xing, which means that the amino acid has been completely connected.

⑥ Cycle: cycle ② - ⑤ repeatedly until synthesis is completed.

⑦ The synthetic peptide is cut and deprotected from the resin. (The Fmoc strategy uses TFA (strong acid) to cut the peptide from the resin.)