RESUMO
Tryptophan 2,3-dioxygenase (TDO) and indoleamine 2,3-dioxygenase (IDO) constitute an important, yet relatively poorly understood, family of heme-containing enzymes. Here, we report extensive structural and biochemical studies of the Xanthomonas campestris TDO and a related protein SO4414 from Shewanella oneidensis, including the structure at 1.6-A resolution of the catalytically active, ferrous form of TDO in a binary complex with the substrate L-Trp. The carboxylate and ammonium moieties of tryptophan are recognized by electrostatic and hydrogen-bonding interactions with the enzyme and a propionate group of the heme, thus defining the L-stereospecificity. A second, possibly allosteric, L-Trp-binding site is present at the tetramer interface. The sixth coordination site of the heme-iron is vacant, providing a dioxygen-binding site that would also involve interactions with the ammonium moiety of L-Trp and the amide nitrogen of a glycine residue. The indole ring is positioned correctly for oxygenation at the C2 and C3 atoms. The active site is fully formed only in the binary complex, and biochemical experiments confirm this induced-fit behavior of the enzyme. The active site is completely devoid of water during catalysis, which is supported by our electrochemical studies showing significant stabilization of the enzyme upon substrate binding.
Assuntos
Triptofano Oxigenase/química , Triptofano Oxigenase/metabolismo , Sítio Alostérico , Sequência de Aminoácidos , Catálise , Cristalografia por Raios X , Humanos , Ligação de Hidrogênio , Técnicas In Vitro , Indolamina-Pirrol 2,3,-Dioxigenase/química , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Cinética , Modelos Moleculares , Dados de Sequência Molecular , Conformação Proteica , Estrutura Quaternária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Homologia de Sequência de Aminoácidos , Shewanella/enzimologia , Shewanella/genética , Eletricidade Estática , Especificidade por Substrato , Triptofano Oxigenase/genética , Xanthomonas campestris/enzimologia , Xanthomonas campestris/genéticaRESUMO
This study describes our experience with an unusual type of acromial spur. In the last 7 years, we have treated 20 patients with a previously undescribed "keeled" acromion. The keel refers to a central, longitudinal, downward sloping spur on the acromial undersurface. Patients with a keeled acromion are at significant risk of bursal-sided as well as full-thickness rotator cuff tears. All patients in this series had significant bursal-sided tears, and 12 of 20 patients (60%) had full-thickness rotator cuff tears associated with an acromial "keel." In addition, patients with a keeled acromion are younger (average age, 52.7 years) than would be expected for patients with rotator cuff disease. Often these patients are misdiagnosed, and effective treatment is delayed. Diagnosis of a keeled acromion relies on a high level of suspicion, as well as knowledge of clinical and radiographic signs consistent with an acromial keel. We discuss the clinical presentation, radiographic findings, associated rotator cuff pathology, as well as methods to avoid intraoperative technical errors when treating patients with an acromial keel. The goal is to raise awareness of this relatively uncommon but clinically important entity and to help guide the treating surgeon's therapeutic measures.
