Characterization of natural and recombinant nuclease isoforms by electrospray mass spectrometry.
Isoforms of natural and recombinant nuclease have been characterized on the basis of their M(r) as determined by electrospray m.s.. The natural nuclease was isolated and purified from Serratia marcescens B10M1 and the recombinant nuclease from Escherichia coli MT102 carrying the plasmid p403-SD2. The primary structure of each of the isoforms isolated from the nuclease preparations was established by comparing their mass with the known amino acid sequence derived from the nucleotide sequence of the nuc gene. All the preparations were found to be contaminated with the same N-terminal split variants of native nuclease, although the natural nuclease contained much larger amounts of these isoforms than did the recombinant nuclease. The structure of some of the isoforms could be verified by N-terminal sequencing, and nearly all of them by isoelectric focusing.
Pedersen
J
J
Department of Biotechnology, Technical University of Denmark, Lyngby.
Andersen
J
J
Roepstorff
P
P
Filimonova
M
M
Biedermann
K
K
D004706:Endodeoxyribonucleases / Q000737:chemistry* / Q000302:isolation & purification; D004722:Endoribonucleases / Q000737:chemistry* / Q000302:isolation & purification; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D007525:Isoelectric Focusing; D007527:Isoenzymes; D013058:Mass Spectrometry; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D012706:Serratia marcescens / Q000201:enzymology*
D007527:Isoenzymes; D011994:Recombinant Proteins; D004706:Endodeoxyribonucleases; D004722:Endoribonucleases; C021835:Serratia marcescens nuclease