The successful engineering of biosynthetic pathways hinges on understanding the factors that influence acyl carrier protein (ACP) stability and function. The stability and structure of ACPs can be influenced by the presence of divalent cations, but how this relates to primary sequence remains poorly understood. As part of a course-based undergraduate research experience, we investigated the thermostability of type II polyketide synthase (PKS) ACPs. We observed an approximate 40 °C range in the thermostability amongst the 14 ACPs studied, as well as an increase in stability (5 – 26 °C) of the ACPs in the presence of divalent cations. Distribution of charges in the helix II-loop-helix III region was found to impact the enthalpy of denaturation. Taken together, our results reveal clues as to how the sequence of type II PKS ACPs relates to their structural stability, information that can be used to study how ACP sequence relates to function.

The placement of caudal epidural catheters in horses has become more frequent as a multi-modal analgesic strategy. Despite its integration into clinical practice, there are limited reports describing the use of caudal epidural catheterization for prolonged use in horses. The purpose of this study was to characterize the hospitalized caseload undergoing epidural catheterization for long-term epidural analgesic administration, to report the response to epidural therapy and observed complications, and to describe patient outcomes. Medical records of hospitalized equine patients that underwent placement of a caudal epidural catheter for analgesic management between 2017 and 2021 were analyzed retrospectively. For the 62 catheters placed in the 48 cases, the most frequent diagnosis category prompting epidural analgesia was orthopedic (43/48, 89.6%). Synovial sepsis was the most frequent specific diagnosis prompting epidural catheter placement (11/48, 22.9%). The initial response to epidural therapy was characterized as positive for 37/62 (59.7%) catheters. Complications were documented for 46/62 (74.2%) catheters. However, most of these complications were classified as mild (51.6%) or moderate (14.5%), and exaggerated physiologic responses were observed most frequently. Of the horses studied, 52.1% survived to be discharged from the hospital. With awareness of potential complications and vigilant monitoring, caudal epidural catheters should be considered for equine patients as an analgesic strategy.

The L-site binding of ferricytochrome c to cardiolipin (CL) -containing liposomes at slightly acidic pH has attracted interest owing to its potential role in converting this classical electron transfer protein into a peroxidase. Various lines of evidence presented by Nantes and coworkers suggest that this mode of binding is electrostatic, active below pH 7, and involves residues K22, K25, and K27 as well as H26 and H33. We combined several spectroscopic techniques to characterize L-site binding thermodynamically and to identify concomitant structural changes. To this end we employed and extended a recently presented thermodynamic model that describes cytochrome c binding to CL-containing liposomes as a two-step process, where native-like liposome-bound conformers convert to more unfolded conformers with an increase in CL concentration. These partially unfolded species are characterized by a loss of the M80 ligand, which is likely replaced by H26 and/or H33. At acidic pH, the partially unfolded low-spin species converts into a mixture of penta-and hexacoordinate high-spin species at moderate-to-high CL concentration. This change of the heme iron's spin state is most likely caused by the protonation of the non-native histidine ligand, which is facilitated by low effective pH at the liposome surface. Results of our analysis of binding isotherms obtained in the absence and presence of NaCl confirmed the electrostatic character of L-site binding. The inhibiting effect of sodium ions is attributed to the accumulation of cations in the liposome's double layer, which reduces its surface potential. Thus, L-site binding is clearly distinct from cytochrome c -CL interactions at neutral pH where only the conformational change on the membrane surface is affected by the presence of NaCl.

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