Purification and Chemical Characterization of β‐Trace Protein from Human Cerebrospinal Fluid: Its Identification as Prostaglandin D Synthase

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Abstract
β‐Trace protein from pooled human CSF was purified to homogeneity. An apparent molecular mass of 23–29 kDa was determined for the polypeptide on sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. Amino‐terminal sequencing of the polypeptide yielded the unique amino acid sequence APEAQVSVQPNFQQDKFLGRWFSA24. Alignment of amino acid sequences obtained from tryptic peptides with the sequence previously deduced from a cDNA clone isolated by other investigators allowed the identification of β‐trace protein as prostaglandin D synthase [prostaglandin‐H2 D‐isomerase; (5Z, 13E)‐(15S)‐9α, 11 a‐epidioxy‐15‐hydroxyprosta‐5,13‐dienoate D‐isomerase; EC 5.3.99.2]. A conservative amino acid exchange (The instead of Ser) was detected at amino acid position 154 of the β‐trace polypeptide chain in the corresponding tryptic peptide. The two N‐glycosylation sites of the polypeptide were shown to be almost quantitatively occupied by carbohydrate. Carbohydrate compositional as well as methylation analysis indicated that Asn29and Asn56 bear exclusively complex‐type oligosaccharide structures (partially sialylated with α2–3‐ and/or α2–6‐linked N‐acetylneuraminic acid) that are almost quantitatively α1‐6 fucosylated at the proximal N‐acetylglucosamine; ∼70% of these molecules contain a bisecting N‐acetylglucosamine. Agalacto structures as well as those with a peripheral fucose are also present.