Kc. Parker et al., IDENTIFICATION OF YEAST PROTEINS FROM 2-DIMENSIONAL GELS - WORKING OUT SPOT CROSS-CONTAMINATION, Electrophoresis, 19(11), 1998, pp. 1920-1932
Citations number
14
Categorie Soggetti
Biochemical Research Methods","Chemistry Analytical
With the complete sequence of the yeast genome now available, efforts
by many laboratories are underway to identify each of the spots on two
-dimensional (2-D) gels corresponding to the most abundant yeast prote
ins. The high mass accuracy now attainable using matrix assisted laser
desorption/ionization (MALDI)-mass spectrometry equipped with delayed
extraction simplifies the process of identification, such that many s
pots can be unambiguously identified in a short period of time merely
by using peptide mass fingerprinting and generally available database
matching programs. Although it is not always possible to match spots b
etween gels run by different laboratories, proteins generally yield th
e same abundant proteolytic fragments when tryptic digestions are perf
ormed. Databases containing these signature peptides not only simplify
the task of reidentifying proteins from different gels, but also make
it possible to identify small amounts of cross-contaminating proteins
from different spots, as well as common extraneous contaminants such
as human keratins. In this paper, we present data on the identificatio
n of > 20 previously unreported yeast proteins from 2-D gels. Some nov
el proteins were identified from randomly analyzed spots. Focusing on
14 spots in a narrow-pH-range gel, we demonstrate how organizing peak-
table data and peptide match-list data into databases enables the iden
tification of a larger percentage of the peaks.