Identification of five hemoglobins in B6C3F1 mice by mass spectrometry and sequence analysis.
The aim of the work is to identify and characterize the hemoglobins found in B6C3F1 mice using mass spectrometry. The primary structures are compared to those reported for BALB/c mice. Individual hemoglobin chains were isolated by reverse-phase high performance liquid chromatography (RP-HPLC). The molecular masses of the globins were determined using electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI). The purified globin chains were enzymatically cleaved and the resulting peptides were separated by RP-HPLC. The chains were identified by N-terminal sequencing and mass spectrometry (MALDI). Selected peptides were analysed by Edman degradation. ESI analysis indicates that B6C3F1 mice have two alpha-globin chains (alpha-1 and alpha-2) and at least three beta-globin chains, beta-1, beta-2 and beta-3. This is one additional alpha- and one additional beta-globin chain than reported in the literature for BALB/c mice. Mass and sequence analysis of enzymatically generated peptides showed variations in the amino acid sequence in the alpha-1, alpha-2, beta-2 and beta-3 chains compared to the BALB/c mouse hemoglobins (alpha, beta (minor) and beta (major)). The study showed that mass spectrometry in combination with traditional protein chemistry is able to identify and locate minor protein sequence variations.
Palm
L
L
NIEHS, Research Triangle Park, NC 27709, USA.
Burka
L T
LT
Højrup
P
P
Stevens
R D
RD
Tomer
K B
KB
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D005914:Globins / Q000235:genetics / Q000302:isolation & purification; D006454:Hemoglobins / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D006454:Hemoglobins; D005914:Globins