2-Acylamido analogues of N-acetylglucosamine prime formation of chitin oligosaccharides by yeast chitin synthase 2.

Chitin, a homopolymer of ß1,4-linked N-acetylglucosamine (GlcNAc) residues, is a key component of the cell walls of fungi and the exoskeletons of arthropods. Chitin synthases transfer GlcNAc from UDP-GlcNAc to preexisting chitin chains in reactions that are typically stimulated by free GlcNAc. The effect of GlcNAc was probed by using a yeast strain expressing a single chitin synthase, Chs2, by examining formation of chitin oligosaccharides (COs) and insoluble chitin, and by replacing GlcNAc with 2-acylamido analogues of GlcNAc. Synthesis of COs was strongly dependent on inclusion of GlcNAc in chitin synthase incubations, and N,N'-diacetylchitobiose (GlcNAc2) was the major reaction product. Formation of both COs and insoluble chitin was also stimulated by GlcNAc2 and by N-propanoyl-, N-butanoyl-, and N-glycolylglucosamine. MALDI analyses of the COs made in the presence of 2-acylamido analogues of GlcNAc showed they that contained a single GlcNAc analogue and one or more additional GlcNAc residues. These results indicate that Chs2 can use certain 2-acylamido analogues of GlcNAc, and likely free GlcNAc and GlcNAc2 as well, as GlcNAc acceptors in a UDP-GlcNAc-dependent glycosyltransfer reaction. Further, formation of modified disaccharides indicates that CSs can transfer single GlcNAc residues.

Synthesis and evaluation of chromogenic and fluorogenic substrates for high-throughput detection of enzymes that hydrolyze inorganic polyphosphate.

Inorganic polyphosphates, linear polymers of orthophosphate, occur naturally throughout biology and have many industrial applications. Their biodegradable nature makes them attractive for a multitude of uses, and it would be important to understand how polyphosphates are turned over enzymatically. Studies of inorganic polyphosphatases are, however, hampered by the lack of high-throughput methods for detecting and quantifying rates of polyphosphate degradation. We now report chromogenic and fluorogenic polyphosphate substrates that permit spectrophotometric monitoring of polyphosphate hydrolysis and allow for high-throughput analyses of both endopolyphosphatase and exopolyphosphatase activities, depending on assay configuration. These substrates contain 4-nitrophenol or 4-methylumbelliferone moieties that are covalently attached to the terminal phosphates of polyphosphate via phosphoester linkages formed during reactions mediated by EDAC (1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide). This report identifies Nudt2 as an inorganic polyphosphatase and also adds to the known coupling chemistry for polyphosphates, permitting facile covalent linkage of alcohols with the terminal phosphates of inorganic polyphosphate.

Estimating the size and scope of the academic emergency physician workforce.

BACKGROUND: Academic emergency medicine (EM) is foundational to the EM specialty through the development of new knowledge and clinical training of resident physicians. Despite recent increased attention to the future of the EM workforce, no evaluations have specifically characterized the U.S. academic EM workforce. We sought to estimate the national proportion of emergency physicians (EPs) identified as academic and the proportion of emergency department (ED) visits that take place at academic sites. METHODS: We performed a cross-sectional analysis of EPs and EDs using data from the American Hospital Association, the Centers for Medicare & Medicaid Services, and Doximity's Residency Navigator. EPs were identified as "academic" if they were affiliated with at least one facility determined to be academic, defined as EDs officially designated by the Accreditation Council for Graduate Medical Education (ACGME) as clinical training sites at accredited EM residency programs. Our primary outcomes were to estimate the national proportion of EPs identified as academic and the proportion of ED visits performed at academic sites. RESULTS: Our analytic sample included 26,937 EPs practicing clinically across 4920 EDs and providing care during 130,471,386 ED visits. Among EPs, 11,720 (43.5%) were identified as academic, and among EDs, 635 (12.9%) were identified as academic sites, including 585 adult/general sites, 45 pediatric-specific sites, and 10 sites affiliated with the Department of Veterans Affairs. In 2021, academic EDs provided care for 42,794,106 ED visits or 32.8% of all ED visits nationally. CONCLUSIONS: Approximately four in 10 EPs practice in at least one clinical training site affiliated with an ACGME-accredited EM residency program, and approximately one in three ED visits nationally occur in these academic EDs. We encourage further work using alternative definitions of an academic EPs and EDs, along with longitudinal research to identify trends in the workforce's composition.