Mass spectrometry captures off-target drug binding and provides mechanistic insights into the human metalloprotease ZMPSTE24.

Off-target binding of hydrophobic drugs can lead to unwanted side effects, either through specific or non-specific binding to unintended membrane protein targets. However, distinguishing the binding of drugs to membrane proteins from that of detergents, lipids and cofactors is challenging. Here, we use high-resolution mass spectrometry to study the effects of HIV protease inhibitors on the human zinc metalloprotease ZMPSTE24. This intramembrane protease plays a major role in converting prelamin A to mature lamin A. We monitored the proteolysis of farnesylated prelamin A peptide by ZMPSTE24 and unexpectedly found retention of the C-terminal peptide product with the enzyme. We also resolved binding of zinc, lipids and HIV protease inhibitors and showed that drug binding blocked prelamin A peptide cleavage and conferred stability to ZMPSTE24. Our results not only have relevance for the progeria-like side effects of certain HIV protease inhibitor drugs, but also highlight new approaches for documenting off-target drug binding

Combining DNP NMR with segmental and specific labeling to study a yeast prion protein strain that is not parallel in-register

The yeast prion protein Sup35NM is a self-propagating amyloid. Despite intense study, there is no consensus on the organization of monomers within Sup35NM fibrils. Some studies point to a â-helical arrangement, whereas others suggest a parallel inregister organization. Intermolecular contacts are often determined by experiments that probe long-range heteronuclear contacts for fibrils templated from a 1:1 mixture of 13 C- and 15 N-labeled monomers. However, for Sup35NM, like many large proteins, chemical shift degeneracy limits the usefulness of this approach. Segmental and specific isotopic labeling reduce degeneracy, but experiments to measure long-range interactions are often too insensitive. To limit degeneracy and increase experimental sensitivity, we combined specific and segmental isotopic labeling schemes with dynamic nuclear polarization (DNP) NMR. Using this combination, we examined an amyloid form of Sup35NM that does not have a parallel in-register structure. The combination of a small number of specific labels with DNP NMR enables determination of architectural information about polymeric protein systems. Keyword: [PSI+] prion; solid-state NMR; amyloid; Sup35; dynamic nuclear polarizationNational Institutes of Health (U.S.) (Grants GM-025874)National Institutes of Health (U.S.) (Grants EB-003151)National Institutes of Health (U.S.) (Grants EB-002804)National Institutes of Health (U.S.) (Grants EB-002026

Use of Exposure Standards in Aviation

The use of occupational exposure limits and threshold limit values in the aircraft environment is examined in relation to aircraft air supplies contaminated by engine oils, hydraulic and other fluids used in aircraft

Aircraft Clean Air Requirements Using Bleed Air Systems

There are certification and airworthiness requirements relevant to the provision of clean breathing air in the crew and passenger compartments. There have been continuing reports and studies over the years regarding oil fumes in aircraft, including impaired crew performance. Oil fumes are viewed in varying ways ranging from rare seal bearing failures, to low level leakage in normal flight. A Masters of Science (MSc) research degree was undertaken to assess whether there is any gap between the certification requirements for the provision of clean air in crew and passenger compartments, and the theoretical and practical implementation of the requirements using the bleed air system. A comprehensive literature search reviewed applicable certification standards, documented and theoretical understanding of oil leakage. Two types of interviews were conducted to address the research questions. Key aviation regulators were questioned about the process by which they certify and ensure compliance with the clean air requirements. Aerospace engineers and sealing professionals were interviewed about their understanding of how oil may leak past compressor oil bearing seals, and into the air supply under various flight conditions. The outcome of the research showed that there is a gap between the clean air certification requirements, and the theoretical and practical implementation of the requirements using the bleed air system. Low level oil leakage into the aircraft cabin in normal flight operations is a function of the design of the engine lubricating system and bleed air systems, both utilising pressurised air. The use of the bleed air system to supply the regulatory required air quality standards is not being met or being enforced as required

Ultrafine particle levels measured on board short-haul commercial passenger jet aircraft

Abstract Background Airline crew members report adverse health effects during and after inhalation exposure to engine oil fumes sourced to the air supply system onboard commercial and military aircraft. Most investigations into the causal factors of their reported symptoms focus on specific chemical contaminants in the fumes. The adverse health effects reported in aircrew exposed to the aircraft air supply, bled unfiltered off the engine or Auxiliary Power Unit (APU) may be related to particulate exposures, which are widely known to effect health. While oil contaminates the aircraft air supply, some suggest that this will only occur when there is a bearing seal failure, others document that there is low level oil contamination of the air supply during normal engine operation. This brief pilot study explores whether particulate exposure may be associated with the normal engine/APU and air supply operation and to therefore increase the understanding that UFP exposures may have on crew and passengers. Methods An ultrafine particle counter was utilised by an experienced airline captain in the passenger cabin of four short-haul commercial passenger aircraft. All flights were under 90 min on aircraft from two different carriers ranging from 7 months to 14 years old. Results UFP concentrations showed maximum concentrations ranging from 31,300 to 97,800 particles/cm3 when APU was selected on as a source of air on the ground and with engine bleed air and the air conditioning packs selected on during the climb. In 2 of the 4 flights the peaks were associated with an engine oil smell. Increases in UFP particle concentrations occurred with changes in engine/APU power and air supply configuration changes. Conclusions This study identified increases in UFP concentrations associated with engine and APU power changes and changes in air supply configuration. These results correlated with times when engine and APU oil seals are known to be less effective, enabling oil leakage to occur. The concentrations reached in the passenger cabins exceeded those taken in other ground-based environments. UFP exposures in aircraft cabins during normal flight indicates there will be health consequences for long serving aircrew and some passengers

Aerotoxic Syndrome: A New Occupational Disease?

Background: Concerns related to adverse health effects experienced by aircrew exposed to aircraft contaminated air have been ongoing for over 6 decades. Unfiltered breathing air is supplied to the cabin via the engine compressor. The likelihood that oil leaking over the engine oil seals may enter the cabin air supply has prompted continuing debate about the hazards associated with exposure to neurotoxic substances and to the thermally degraded or pyrolysed mixture. In this study, we undertook an in-depth investigation of aircrew involved in suspected aircraft contaminated air events.  Methods: Two studies were conducted to review the circumstances and symptoms of a cohort of aircrew working in the pressurized air environment of aircraft. A table of effects was then used for categorizing symptoms and reviewing other sources of data related to aircraft fluids and selected other conditions.  Results: Both acute and chronic exposures to neurotoxic and a wide range of thermally degraded substances were confirmed, along with a clear pattern of acute and chronic adverse effects. The latter were supported by medical findings and diagnoses, notably involving the neurological, neurobehavioural and respiratory systems.  Conclusion: A clear cause and effect relationship has been identified linking the symptoms, diagnoses and findings to the occupational environment. Recognition of this new occupational disorder and a clear medical investigation protocol are urgently needed