PMID- 2164192
OWN - NLM
STAT- MEDLINE
DCOM- 19900814
LR  - 20190818
IS  - 0724-8741 (Print)
IS  - 0724-8741 (Linking)
VI  - 7
IP  - 6
DP  - 1990 Jun
TI  - Chemical pathways of peptide degradation. I. Deamidation of adrenocorticotropic 
      hormone.
PG  - 593-9
AB  - Deamidation of Asn residues is one of the major chemical pathways of degradation 
      of proteins and peptides. Adrenocorticotropic hormone (ACTH), a 39-amino acid 
      polypeptide with a single Asn residue, was shown in this study to be a useful 
      model polypeptide for the study of the effects of pH and buffer concentration on 
      the rate and pathway of deamidation. The disappearance of ACTH and appearance of 
      deamidated ACTH were monitored by isoelectric focusing (IEF), and ammonia 
      production was monitored spectrophotometrically using a coupled enzymatic assay. 
      Using these analytical methods, the deamidation of ACTH was shown to follow 
      pseudo-first-order kinetics and was dependent on pH and buffer concentrations. 
      The separation of the deamidated ACTHs (Asp-ACTH and isoAsp-ACTH) from ACTH was 
      successful, but attempts to separate Asp-ACTH from isoAsp-ACTH using 
      cation-exchange HPLC and IEF were unsuccessful. Using bovine protein 
      carboxymethyltransferase (PCM), which selectively methylates the carboxyl group 
      of isoAsp residue, the isoAsp-ACTH could be detected at pH 7.0 and 9.6 but not at 
      pH 1.9. These data support the hypothesis that under neutral and alkaline 
      conditions, deamidation of ACTH proceeds through the formation of a cyclic imide 
      intermediate (slow step), followed by its hydrolysis to the Asp-ACTH and 
      isoAsp-ACTH (fast step). Under acidic conditions, the reaction appears to proceed 
      via direct hydrolysis of the Asn residue to form Asp-ACTH without the formation 
      of a cyclic imide intermediate.
FAU - Bhatt, N P
AU  - Bhatt NP
AD  - Department of Pharmaceutical Chemistry, University of Kansas, Lawrence 66045.
FAU - Patel, K
AU  - Patel K
FAU - Borchardt, R T
AU  - Borchardt RT
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Pharm Res
JT  - Pharmaceutical research
JID - 8406521
RN  - 0 (Amides)
RN  - 0 (Buffers)
RN  - 0 (Indicators and Reagents)
RN  - 0 (Peptides)
RN  - 7664-41-7 (Ammonia)
RN  - 7LP2MPO46S (S-Adenosylmethionine)
RN  - 9002-60-2 (Adrenocorticotropic Hormone)
RN  - EC 2.1.1.- (Protein O-Methyltransferase)
SB  - IM
MH  - Adrenocorticotropic Hormone/*analysis
MH  - Amides/analysis
MH  - Ammonia/analysis
MH  - Buffers
MH  - Chromatography, High Pressure Liquid
MH  - Hydrogen-Ion Concentration
MH  - Indicators and Reagents
MH  - Ion Exchange
MH  - Isoelectric Focusing
MH  - Kinetics
MH  - Peptides/isolation & purification
MH  - Protein O-Methyltransferase/analysis
MH  - S-Adenosylmethionine/analysis
EDAT- 1990/06/01 00:00
MHDA- 1990/06/01 00:01
CRDT- 1990/06/01 00:00
PHST- 1990/06/01 00:00 [pubmed]
PHST- 1990/06/01 00:01 [medline]
PHST- 1990/06/01 00:00 [entrez]
AID - 10.1023/a:1015862026539 [doi]
PST - ppublish
SO  - Pharm Res. 1990 Jun;7(6):593-9. doi: 10.1023/a:1015862026539.