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111.
Daniel L. Hofmeister Chase A. Seltzner Nicholas J. Bockhaus James B. Thoden Hazel M. Holden 《Protein science : a publication of the Protein Society》2023,32(1):e4502
Psychrobacter cryohalolentis K5T is a Gram-negative bacterium first isolated from Siberian permafrost in 2006. It has a complex O-antigen containing l -rhamnose, d -galactose, two diacetamido-sugars, and one triacetamido-sugar. The biosynthetic pathway for one of the diacetamido-sugars, namely 2,3-diacetamido-2,3-dideoxy-d -glucuronic acid, is presently unknown. Utilizing the published genome sequence of P. cryohalolentis K5T, we hypothesized that the genes designated Pcryo_0613, Pcryo_0614, Pcryo_0616, and Pcryo_0615 encode for a uridine dinucleotide (UDP)-N-acetyl-d -glucosamine 6-dehydrogenase, an nicotinamide adenine dinucleotide (oxidized) (NAD+)-dependent dehydrogenase, a pyridoxal 5′-phosphate (PLP)-dependent aminotransferase, and an N-acetyltransferase, respectively, activities of which would be required for the biosynthesis of this unusual carbohydrate. Here we present the cloning, overexpression, and purification of these hypothetical proteins. Kinetic data on the enzymes encoded by Pcryo_0613, Pcryo_0614, and Pcryo_0615 confirmed their postulated biochemical activities. In addition, the high-resolution X-ray structures of both the internal and external aldimine forms of the aminotransferase were determined to 1.25 and 1.0 Å, respectively. Finally, the three-dimensional architecture of the N-acetyltransferase in complex with its substrate and coenzyme A was solved to 1.8 Å resolution. Strikingly, the N-acetyltransferase was shown to adopt a new motif for UDP-sugar binding. The data presented herein provide additional insight into sugar biosynthesis in Gram-negative bacteria. 相似文献
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I Shelub A van Grondelle R McCullough S Hofmeister J T Reeves 《Journal of applied physiology (Bethesda, Md. : 1985)》1984,56(3):810-815
Despite numerous efforts, a reliable model of chronic embolic pulmonary hypertension has not been established. To develop such a model five conscious mongrel dogs were embolized repeatedly over 16-30 wk with Sephadex microspheres 286 +/- 70 micron in diameter. Hemodynamic and respiratory measurements were obtained just prior to each embolization. Chronic pulmonary hypertension developed in all dogs. Pulmonary hypertension was not accounted for by increased cardiac output, wedge pressure, right atrial pressure, or systemic arterial pressure. Gas exchange was little altered. Lung histological study revealed microspheres clustered within vessels. In three dogs increased pulmonary arterial pressure was sustained despite cessation of embolization for up to 5 mo. Reembolization in one of these caused further pulmonary hypertension. In two dogs acute pulmonary vasodilation by O2 breathing and administration of prostaglandin E1 reduced, but did not abolish, the increased pulmonary vascular resistance, suggesting some vascular tone was present. An embolic model of chronic pulmonary hypertension in awake dogs allows further investigation into the evolution of pulmonary hypertension. 相似文献