Internal Protein Sequence
Research Committee
The ABRF Internal Protein Sequence Research Committee was formed under the
Protein Sequence Research Committee for the purpose of evaluating current
methods available for obtaining sequences other than the amino-terminus of
proteins. Six laboratories are participating in this subcommittee, which is
using different methods for obtaining internal protein sequence data from
SDS-PAGE gels, nitrocellulose, and PVDF samples. Standard proteins (human
transferrin, carbonic anhydrase, bovine serum albumin, b-galactosidase,
ovalbumin) in varying quantities (10-50 pmol) were enzymatically digested
using various enzymes. For each study identical protein samples were
prepared by the Wistar Laboratory as SDS-PAGE gel, nitrocellulose, or PVDF
samples and distributed to the participating laboratories for digestion and
elution. Standard proteins in solution and appropriate blanks were also
digested as controls. All digestions were sent to the Rockefeller
University for HPLC analysis using an HP 1090 binary solvent system
equipped with a diode array detector, autosampler, and Vydac C18 column
(2.1 x 250 mm).
The participating laboratories are Ryuji Kobayashi and Nora Poppito (Cold
Spring Harbor Laboratory), Bill Lane and Renee Robinson (Harvard
University), Paul Tempst and Hediye Erdjument-Bromage (Memorial Sloan
Kettering Cancer Center), Sheenah Mische and Joe Fernandez (Rockefeller
University), Dave Speicher and Dave Reim (Wistar Institute), and Ken
Williams and Kathy Stone (Yale University). The preliminary results of this
study will be presented as a poster at the 1995 ABRF Meeting in Boston.
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Created: 27th July 1995
Last modified: 27th July 1995