Structural comparison of bovine erythrocyte, brain, and liver NADH-cytochrome b5 reductase by HPLC mapping.
NADH-cytochrome b5 reductases purified from bovine erythrocytes and from bovine brain and liver microsomes solubilized with lysosomal protease were subjected to structural analysis by using HPLC mapping, amino acid analysis of the resulting peptides, and NH2-terminal sequence analysis of apoproteins. HPLC maps of the tryptic peptides derived from these enzymes were very similar to each other, and amino acid analysis of the HPLC-separated peptides indicated that the structures of these enzymes are identical except for the NH2-terminal region. The NH2-terminal sequence of the brain enzyme determined by automated Edman degradation was as follows: NH2-Phe-Gln-Arg-Ser-Thr-Pro-Ala-Ile-Thr-Leu-Glu-Asn-Pro-Asp- Ile-Lys-Tyr-Pro-Leu-Arg-Leu-Ile-Asp-Lys-Glu-Val-Ile- This sequence is identical to that of liver enzyme except that the liver enzyme started at the 3rd Arg or 4th Ser. The NH2-terminal amino acid residue of the soluble erythrocyte enzyme was not detected by automated Edman degradation. The sequence analysis of a tryptic peptide from the erythrocyte enzyme indicated that Leu is present before the NH2-terminal Phe of the brain enzyme. The recently reported sequence of the apparently identical protein (Ozols et al. (1985) J. Biol. Chem. 260, 11953-11961) differs in two amino acid assignments from our sequence.
Tamura
M
M
Department of Biochemistry, Medical College of Oita.
Yubisui
T
T
Takeshita
M
M
Kawabata
S
S
Miyata
T
T
Iwanaga
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003579:Cytochrome Reductases / Q000097:blood / Q000302:isolation & purification*; D042966:Cytochrome-B(5) Reductase; D004912:Erythrocytes / Q000201:enzymology*; D008099:Liver / Q000201:enzymology*; D009928:Organ Specificity; D010449:Peptide Mapping
D003579:Cytochrome Reductases; D042966:Cytochrome-B(5) Reductase