PAM and MBHA resins can be used with the Boc chemistry on the ABI 433A instrument.
Each resin provides a unique feature to the final product. • PAM resins—a carboxylic acid terminal peptide • MBHA resins—an amide terminal peptide
PAM Resins
PAM resins were developed to minimize the loss of peptide chains during SPPS.35
The name “PAM” is derived from the linker, 4-(oxymethyl)- phenylacetamidomethyl. A PAM resin structure is shown in Figure 3-21.
Figure 3-21. PAM resin
With the conventional Merrifield-type resin, up to 0.7% of the peptide can be lost with each TFA treatment. In comparison, with the PAM resin only 0.007% loss occurs per cycle.36
Operations and Reagents 0.50 mmol 0.10 mmol
TFA deprotection
30% TFA in DCM 3 3
50% TFA in DCM 16 11
Washes and neutralizations
DCM washes (5X) 3 2
5% DIEA 4 2
NMP washes (6X) 5 3
Coupling
Boc-AA-HOBt ester in NMP 39 23
DMSO to make 15% DMSO/85% NMP 16 8
3.8 equiv. DIEA 5 4
Wash and resin sample NMP wash
Resin sample (optional) (2) (2)
Capping
10% Ac2O, 5% DIEA in NMP 9 5
Washes
DCM washes (6X) 4 4
————— —————
Total Cycle Time (without resin sample) 104 65
N H CH R C O O CH2 Boc CH2 C O NH CH2 Resin
Besides increased acid stability, properly synthesized PAM resins offer two additional advantages: they contain few aldehydes and no extra hydroxy- methyl sites. Aldehyde functionalities, when found in conventional resins, can cause Schiff’s base formation with the peptide amine, resulting in a deletion peptide.37
Furthermore, conventional resins may have
hydroxymethyl sites which, through trifluoroacetylation, can cause chain termination.38
MBHA Resins
The structure of MBHA (methylbenzhydrylamine) resin in the free amine form is shown in Figure 3-22. Even though it may not be indicated on the label, it is always purchased as the hydrochloride salt.
The first amino acid can easily be attached to the MBHA resin using the normal coupling cycle. Cleavage with a strong acid (such as HF or TFMSA) is necessary to remove the amide peptide from MBHA resins.
Figure 3-22. MBHA resin
A clever use of MBHA resins incorporates Boc-Asp(α-OBzl) or Boc-Glu(α- OBzl) anchored to MBHA resin by its side chain. After cleavage with HF or TFMSA, the peptide contains C-terminal asparagine or glutamine.38
Cleavage
The booklet entitled Introduction to Cleavage Techniques presents a compre- hensive discussion of cleavage procedures and scavengers for both Fmoc and Boc chemistry.
This booklet is available on the web: www.appliedbiosystems.com > Service
and Support > Product & Service Literature and enter 343901 in the Docu- ment title box. It is 71 pages long. A shorter reference “Cleavage, Deprotec- tion and Isolation of peptides after Fmoc synthesis” can be obtained on the same web site using document number 123507 and is 12 pages long. Although the peptide resins obtained with FastMoc chemistry are not specif- ically discussed in the booklet, they are Fmoc-peptide resins. You can use the same cleavage techniques on the products of FastMoc chemistry as those used with the products of Fmoc chemistry with HOBt/DCC.
Note When you use the derivatives Fmoc-Asn(Trt) or Fmoc-Gln(Trt),
with more than 0.5 g of peptide resin, use twice the amount of cleavage mixture recommended.
Resin CH
NH2 CH3
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