Isolation and primary structure of NGAL, a novel protein associated with human neutrophil gelatinase.
A 25-kDa protein was found to be associated with purified human neutrophil gelatinase. Polyclonal antibodies raised against gelatinase not only recognized gelatinase but also this 25-kDa protein. Specific antibodies against the 25-kDa protein were obtained by affinity purification of the gelatinase antibodies. Immunoblotting and immunoprecipitation studies demonstrated the 135-kDa form of gelatinase to be a complex of 92-kDa gelatinase and the 25-kDa protein, and the 220-kDa form was demonstrated to be a homodimer of the 92-kDa protein, thus explaining the 220-, 135-, and 92-kDa forms characteristic of neutrophil gelatinase. The 25-kDa protein was purified to apparent homogeneity from exocytosed material from phorbol myristate acetate-stimulated neutrophils. The primary structure of the 25-kDa protein was determined as a 178-residue protein. It was susceptible to treatment with N-glycanase, and one N-glycosylation site was identified. The sequence did not match any known human protein, but showed a high degree of similarity with the deduced sequences of rat alpha 2-microglobulin-related protein and the mouse protein 24p3. It is thus a new member of the lipocalin family. The function of the 25-kDa protein, named neutrophil gelatinase-associated lipocalin (NGAL), remains to be determined.
Kjeldsen
L
L
Departments of Hematology, University Hospital, Rigshospitalet, Copenhagen, Denmark.
Johnsen
A H
AH
Sengeløv
H
H
Borregaard
N
N
D000209:Acute-Phase Proteins; D000510:Alpha-Globulins / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D015153:Blotting, Western; D002352:Carrier Proteins / Q000097:blood* / Q000737:chemistry / Q000302:isolation & purification*; D002853:Chromatography, Liquid; D017364:Collagenases / Q000737:chemistry / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000097:blood* / Q000302:isolation & purification*; D006026:Glycoside Hydrolases; D006801:Humans; D000071068:Lipocalin-2; D054834:Lipocalins; D046911:Macromolecular Substances; D020780:Matrix Metalloproteinase 9; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology*; D015513:Oncogene Proteins; D011518:Proto-Oncogene Proteins; D051381:Rats; D017386:Sequence Homology, Amino Acid; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000209:Acute-Phase Proteins; D000510:Alpha-Globulins; D000906:Antibodies; D002352:Carrier Proteins; D006023:Glycoproteins; C489506:LCN2 protein, human; D000071068:Lipocalin-2; D054834:Lipocalins; D046911:Macromolecular Substances; D015513:Oncogene Proteins; D011518:Proto-Oncogene Proteins; C025500:alpha(2)-microglobulin; D006026:Glycoside Hydrolases; C020668:glycanase; D017364:Collagenases; D020780:Matrix Metalloproteinase 9; D013755:Tetradecanoylphorbol Acetate