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A Fluorescence-Based High-Throughput Coupled Enzymatic Assay for Quantitation of Isoaspartate in Proteins and Peptides.

Abstract Formation of isoaspartate (IsoAsp) from spontaneous asparagine (Asn) deamidation or aspartate (Asp) isomerization is one of the most common non-enzymatic pathways of chemical degradation of protein and peptide pharmaceuticals. Rapid quantitation of IsoAsp formation can enable rank-ordering of potential drug candidates, mutants, and formulations as well as support shelf life prediction and stability requirements. A coupled enzymatic fluorescence-based IsoAsp assay (CEFIA) was developed as a high-throughput method for quantitation of IsoAsp in peptides and proteins. In this note, application of this method to two therapeutic candidate proteins with distinct structural scaffolds is described. In addition, the results obtained with this method are compared to those from conventional assays.
PMID
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Authors

Mayor MeshTerms
Keywords

deamidation

fluorescence

high-throughput

isoaspartate

protein

stability

Journal Title aaps pharmscitech
Publication Year Start
%A Puri, Aastha; Quan, Yong; Narang, Ajit S.; Adams, Monica; Gandhi, Rajesh; Nashine, Vishal C.
%T A Fluorescence-Based High-Throughput Coupled Enzymatic Assay for Quantitation of Isoaspartate in Proteins and Peptides.
%J AAPS PharmSciTech
%D 06/2016
%M ENG
%B Formation of isoaspartate (IsoAsp) from spontaneous asparagine (Asn) deamidation or aspartate (Asp) isomerization is one of the most common non-enzymatic pathways of chemical degradation of protein and peptide pharmaceuticals. Rapid quantitation of IsoAsp formation can enable rank-ordering of potential drug candidates, mutants, and formulations as well as support shelf life prediction and stability requirements. A coupled enzymatic fluorescence-based IsoAsp assay (CEFIA) was developed as a high-throughput method for quantitation of IsoAsp in peptides and proteins. In this note, application of this method to two therapeutic candidate proteins with distinct structural scaffolds is described. In addition, the results obtained with this method are compared to those from conventional assays.
%Y 10.1208/s12249-016-0570-7
%W PHY
%G AUTHOR
%R 2016..............P

@Article{Puri2016,
author="Puri, Aastha
and Quan, Yong
and Narang, Ajit S.
and Adams, Monica
and Gandhi, Rajesh
and Nashine, Vishal C.",
title="A Fluorescence-Based High-Throughput Coupled Enzymatic Assay for Quantitation of Isoaspartate in Proteins and Peptides.",
journal="AAPS PharmSciTech",
year="2016",
month="Jun",
day="24",
abstract="Formation of isoaspartate (IsoAsp) from spontaneous asparagine (Asn) deamidation or aspartate (Asp) isomerization is one of the most common non-enzymatic pathways of chemical degradation of protein and peptide pharmaceuticals. Rapid quantitation of IsoAsp formation can enable rank-ordering of potential drug candidates, mutants, and formulations as well as support shelf life prediction and stability requirements. A coupled enzymatic fluorescence-based IsoAsp assay (CEFIA) was developed as a high-throughput method for quantitation of IsoAsp in peptides and proteins. In this note, application of this method to two therapeutic candidate proteins with distinct structural scaffolds is described. In addition, the results obtained with this method are compared to those from conventional assays.",
issn="1530-9932",
doi="10.1208/s12249-016-0570-7",
url="http://www.ncbi.nlm.nih.gov/pubmed/27342117",
language="ENG"
}

%0 Journal Article
%T A Fluorescence-Based High-Throughput Coupled Enzymatic Assay for Quantitation of Isoaspartate in Proteins and Peptides.
%A Puri, Aastha
%A Quan, Yong
%A Narang, Ajit S.
%A Adams, Monica
%A Gandhi, Rajesh
%A Nashine, Vishal C.
%J AAPS PharmSciTech
%D 2016
%8 Jun 24
%@ 1530-9932
%G ENG
%F Puri2016
%X Formation of isoaspartate (IsoAsp) from spontaneous asparagine (Asn) deamidation or aspartate (Asp) isomerization is one of the most common non-enzymatic pathways of chemical degradation of protein and peptide pharmaceuticals. Rapid quantitation of IsoAsp formation can enable rank-ordering of potential drug candidates, mutants, and formulations as well as support shelf life prediction and stability requirements. A coupled enzymatic fluorescence-based IsoAsp assay (CEFIA) was developed as a high-throughput method for quantitation of IsoAsp in peptides and proteins. In this note, application of this method to two therapeutic candidate proteins with distinct structural scaffolds is described. In addition, the results obtained with this method are compared to those from conventional assays.
%U http://dx.doi.org/10.1208/s12249-016-0570-7
%U http://www.ncbi.nlm.nih.gov/pubmed/27342117

PT Journal
AU Puri, A
   Quan, Y
   Narang, AS
   Adams, M
   Gandhi, R
   Nashine, VC
TI A Fluorescence-Based High-Throughput Coupled Enzymatic Assay for Quantitation of Isoaspartate in Proteins and Peptides.
SO AAPS PharmSciTech
PD Jun
PY 2016
DI 10.1208/s12249-016-0570-7
LA ENG
AB Formation of isoaspartate (IsoAsp) from spontaneous asparagine (Asn) deamidation or aspartate (Asp) isomerization is one of the most common non-enzymatic pathways of chemical degradation of protein and peptide pharmaceuticals. Rapid quantitation of IsoAsp formation can enable rank-ordering of potential drug candidates, mutants, and formulations as well as support shelf life prediction and stability requirements. A coupled enzymatic fluorescence-based IsoAsp assay (CEFIA) was developed as a high-throughput method for quantitation of IsoAsp in peptides and proteins. In this note, application of this method to two therapeutic candidate proteins with distinct structural scaffolds is described. In addition, the results obtained with this method are compared to those from conventional assays.
ER

PMID- 27342117
OWN - NLM
STAT- Publisher
DA  - 20160625
LR  - 20160625
IS  - 1530-9932 (Electronic)
IS  - 1530-9932 (Linking)
DP  - 2016 Jun 24
TI  - A Fluorescence-Based High-Throughput Coupled Enzymatic Assay for Quantitation of 
      Isoaspartate in Proteins and Peptides.
AB  - Formation of isoaspartate (IsoAsp) from spontaneous asparagine (Asn) deamidation 
      or aspartate (Asp) isomerization is one of the most common non-enzymatic pathways
      of chemical degradation of protein and peptide pharmaceuticals. Rapid
      quantitation of IsoAsp formation can enable rank-ordering of potential drug
      candidates, mutants, and formulations as well as support shelf life prediction
      and stability requirements. A coupled enzymatic fluorescence-based IsoAsp assay
      (CEFIA) was developed as a high-throughput method for quantitation of IsoAsp in
      peptides and proteins. In this note, application of this method to two
      therapeutic candidate proteins with distinct structural scaffolds is described.
      In addition, the results obtained with this method are compared to those from
      conventional assays.
FAU - Puri, Aastha
AU  - Puri A
AD  - Drug Product Science & Technology, Bristol-Myers Squibb, Co., 1 Squibb Drive, New
      Brunswick, New Jersey, 08901, USA.
FAU - Quan, Yong
AU  - Quan Y
AD  - Drug Product Science & Technology, Bristol-Myers Squibb, Co., 1 Squibb Drive, New
      Brunswick, New Jersey, 08901, USA.
FAU - Narang, Ajit S
AU  - Narang AS
AD  - Drug Product Science & Technology, Bristol-Myers Squibb, Co., 1 Squibb Drive, New
      Brunswick, New Jersey, 08901, USA.
FAU - Adams, Monica
AU  - Adams M
AD  - Drug Product Science & Technology, Bristol-Myers Squibb, Co., 1 Squibb Drive, New
      Brunswick, New Jersey, 08901, USA.
FAU - Gandhi, Rajesh
AU  - Gandhi R
AD  - Drug Product Science & Technology, Bristol-Myers Squibb, Co., 1 Squibb Drive, New
      Brunswick, New Jersey, 08901, USA.
FAU - Nashine, Vishal C
AU  - Nashine VC
AD  - Drug Product Science & Technology, Bristol-Myers Squibb, Co., 1 Squibb Drive, New
      Brunswick, New Jersey, 08901, USA. [email protected]
LA  - ENG
PT  - JOURNAL ARTICLE
DEP - 20160624
TA  - AAPS PharmSciTech
JT  - AAPS PharmSciTech
JID - 100960111
OTO - NOTNLM
OT  - deamidation
OT  - fluorescence
OT  - high-throughput
OT  - isoaspartate
OT  - protein
OT  - stability
EDAT- 2016/06/28 06:00
MHDA- 2016/06/28 06:00
CRDT- 2016/06/26 06:00
PHST- 2016/03/11 [received]
PHST- 2016/06/07 [accepted]
PHST- 2016/06/24 [aheadofprint]
AID - 10.1208/s12249-016-0570-7 [doi]
AID - 10.1208/s12249-016-0570-7 [pii]
PST - aheadofprint
SO  - AAPS PharmSciTech. 2016 Jun 24.
TY  - JOUR
AU  - Puri, Aastha
AU  - Quan, Yong
AU  - Narang, Ajit S.
AU  - Adams, Monica
AU  - Gandhi, Rajesh
AU  - Nashine, Vishal C.
PY  - 2016/Jun/24
TI  - A Fluorescence-Based High-Throughput Coupled Enzymatic Assay for Quantitation of Isoaspartate in Proteins and Peptides.
JO  - AAPS PharmSciTech
N2  - Formation of isoaspartate (IsoAsp) from spontaneous asparagine (Asn) deamidation or aspartate (Asp) isomerization is one of the most common non-enzymatic pathways of chemical degradation of protein and peptide pharmaceuticals. Rapid quantitation of IsoAsp formation can enable rank-ordering of potential drug candidates, mutants, and formulations as well as support shelf life prediction and stability requirements. A coupled enzymatic fluorescence-based IsoAsp assay (CEFIA) was developed as a high-throughput method for quantitation of IsoAsp in peptides and proteins. In this note, application of this method to two therapeutic candidate proteins with distinct structural scaffolds is described. In addition, the results obtained with this method are compared to those from conventional assays.
SN  - 1530-9932
UR  - http://dx.doi.org/10.1208/s12249-016-0570-7
UR  - http://www.ncbi.nlm.nih.gov/pubmed/27342117
ID  - Puri2016
ER  - 
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