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A serine esterase released by human alveolar macrophages is closely related to liver microsomal carboxylesterases.
Munger, J S; Shi, G P; Mark, E A; Chin, D T; Gerard, C; Chapman, H A.
Afiliação
  • Munger JS; Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02115.
J Biol Chem ; 266(28): 18832-8, 1991 Oct 05.
Article em En | MEDLINE | ID: mdl-1918003
We identified a 60-kDa diisopropylfluorophosphate-(DFP) reactive protein in human bronchoalveolar lavage fluid, at a yield of 50-100 pmol/lavage. The protein is associated with the cell-free lavage fluid sediment, which consists mainly of surfactant. [3H]DFP labeling is inhibited by heating to 56 degrees C, 2 mM phenylmethylsulfonylfluoride and 1 mM bis(4-nitrophenyl)-phosphate. An identical 60-kDa [3H]DFP-reactive protein is present in the insoluble fraction of alveolar macrophage-conditioned culture medium and in total membrane preparations of alveolar macrophages. The [3H]DFP-labeled protein was purified approximately 30-fold from lavage fluid sediment by size-exclusion (Sephacryl S-200) and ion-exchange (Mono-Q) chromatography. Cyanogen bromide treatment of the partially purified protein produced a major labeled peptide of 14 kDa with an NH2-terminal sequence 90% identical to a region of form 1 rabbit liver microsomal carboxylesterase. Esterase activity in unlabeled starting material, detected using p-nitrophenyl valerate as substrate, copurified with the [3H]DFP-labeled enzyme. Degenerate oligonucleotide primers were designed based on the partial amino acid sequence and on a highly conserved region of known liver carboxylesterase sequences. Polymerase chain reaction using these primers and reverse-transcribed human alveolar macrophage mRNA yielded a 354-base pair product which was then used to screen a human alveolar macrophage cDNA library. A complete esterase sequence was obtained from two incomplete, overlapping clones, and is virtually identical to human liver carboxylesterase partial sequences. Northern blot analysis demonstrated a single approximately 1.7-kilobase transcript in human monocytes and alveolar macrophages, with much higher levels in the latter. These data indicate that human alveolar macrophages both contain and release a serine esterase that is apparently identical to liver microsomal carboxylesterase. Its enzymatic profile suggests it is a major component of alveolar macrophage-nonspecific esterase activity. We hypothesize that it acts as a detoxication enzyme in the lung.
Assuntos
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Bases de dados: MEDLINE Assunto principal: Microssomos Hepáticos / Hidrolases de Éster Carboxílico / Esterases / Macrófagos Limite: Adult / Humans Idioma: En Revista: J Biol Chem Ano de publicação: 1991 Tipo de documento: Article
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Bases de dados: MEDLINE Assunto principal: Microssomos Hepáticos / Hidrolases de Éster Carboxílico / Esterases / Macrófagos Limite: Adult / Humans Idioma: En Revista: J Biol Chem Ano de publicação: 1991 Tipo de documento: Article