Vibrational reporters have shown significant promise as sensitive probes of local environments in proteins and nucleic acids. The utility of two potential vibrational probes, the cyanate and azide groups in phenyl cyanate and 3-azidopyridine, respectively, has been hindered by accidental Fermi resonance. Anharmonic coupling, between the fundamental –OCN or –N3 asymmetric stretch vibration with a near resonant combination band, results in an extremely broad and complex absorption profile for each of these probes. A total of eight phenyl cyanate and six 3-azidopyridine isotopomers were synthesized and studied. Isotopic editing effectively modulated the accidental Fermi resonance — the absorption profiles of several isotopomers were greatly simplified while others remained complex. The origins of the observed profiles are discussed. Addition of a single neutron to the middle atom of the oscillator converted the absorption profile to essentially a single band resulting from either the cyanate or azide asymmetric stretch vibration.

The ability to genetically incorporate amino acids modified with spectroscopic reporters site-specifically into proteins with high efficiency and fidelity has greatly enhanced the ability to probe local protein structure and dynamics. Here, we have synthesized the unnatural amino acid (UAA), 4-cyano-L-phenylalanine (pCNPhe), containing the nitrile vibrational reporter and three isotopomers ((15)N, (13)C, (13)C(15)N) of this UAA to enhance the ability of pCNPhe to study local protein environments. Each pCNPhe isotopic variant was genetically incorporated in an efficient, site-specific manner into superfolder green fluorescent protein (sfGFP) in response to an amber codon with high fidelity utilizing an engineered, orthogonal aminoacyl-tRNA synthetase. The isotopomers of 4-cyano-L-phenylalanine permitted the nitrile symmetric stretch vibration of these UAAs to be unambiguously assigned utilizing the magnitude and direction of the isotopic shift of this vibration. The sensitivity of the nitrile symmetric stretching frequency of each isotopic variant to the local environment was measured by individually incorporating the probes into two distinct local environments of sfGFP. The UAAs were also utilized in concert to probe multiple local environments in sfGFP simultaneously to increase the utility of 4-cyano-L-phenylalanine.

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Background: Pancreatic duct stents are frequently placed for prophylaxis of post-endoscopic retrograde cholangiopancreatography pancreatitis. Because of concern for possible secondary ductal changes from a retained stent, these stents need to be monitored and removed if retained. Usually an abdominal X-ray is performed to assess retained stent, and if present, an esophagogastroduodenoscopy is performed to remove the stent. Limited data is published on false-negative radiology reports for spontaneous passage of stents. Methods: Using an Institutional Review Board–approved stent log, a retrospective chart review of all pancreatic duct stents placed at our institution from 2008 to 2014 was performed. Results: A total of 856 pancreatic duct stents were placed during the study period. Of these, 435 (50.8%) were prophylactic stents and 421 (49.2%) were therapeutic. Complete follow-up data were available in 426 (97.9%) patients with prophylactic stents. Six patients (1.4%) were lost to follow up and three (0.7%) expired prior to removal. In all, 283 (66%) had follow-up imaging, with 167 (39.2%) having the official radiology read with no retained pancreatic duct stent in place. Eight of these cases were “false-negative” radiology interpretation (4.8% of cases read as “no stent,” NNH = 20). The stent was found either by review of image by an endoscopist or incidental stent discovery during a follow-up procedure. Conclusion: Radiologist interpretation of abdominal X-rays to assess spontaneous passage of prophylactic pancreatic ducts stents resulted in a false-negative interpretation in approximately 5% of cases. Independent review of the images by the endoscopist may be beneficial given unfamiliarity of these stents by radiologists.