Folding mechanism of the metastable serpin α1-antitrypsin.

The misfolding of serpins is linked to several genetic disorders including emphysema, thrombosis, and dementia. During folding, inhibitory serpins are kinetically trapped in a metastable state in which a stretch of residues near the C terminus of the molecule are exposed to solvent as a flexible loop (the reactive center loop). When they inhibit target proteases, serpins transition to a stable state in which the reactive center loop forms part of a six-stranded ß-sheet. Here, we use hydrogen-deuterium exchange mass spectrometry to monitor region-specific folding of the canonical serpin human α(1)-antitrypsin (α(1)-AT). We find large differences in the folding kinetics of different regions. A key region in the metastable → stable transition, ß-strand 5A, shows a lag phase of nearly 350 s. In contrast, the "B-C barrel" region shows no lag phase and the incorporation of the C-terminal residues into ß-sheets B and C is largely complete before the center of ß-sheet A begins to fold. We propose this as the mechanism for trapping α(1)-AT in a metastable form. Additionally, this separation of timescales in the folding of different regions suggests a mechanism by which α(1)-AT avoids polymerization during folding.

Sevoflurane and renal function: a meta-analysis of randomized trials.

OBJECTIVE: This study aims to describe the overall cumulative effect of sevoflurane on kidney function in healthy patients in terms of mean plasma creatinine, blood urea nitrogen (BUN), creatinine clearance, urinary protein, and glucose excretion at 24 and 72 hours post-anesthesia. DATA RETRIEVAL: A systematic literature search using MEDLINE and EMBASE as primary search engines was conducted. Articles, relevant abstracts, and citations dated January 1, 1995 to June 30, 2016 were retrieved. DATA SELECTION: Search terms included the pharmacological generic name sevoflurane. Search was expanded using the terms "renal function" OR "kidney" function AND "creatinine" OR "blood urea nitrogen" OR "creatinine clearance" OR "proteinuria" OR "glucosuria" OR "nephrotoxicity." Limitations included randomized controlled trial, humans, and ages 19 and above, to include English and non-English text formats. All bibliographic indices for the relevant journals identified were also searched and collated according to relevance. MAIN OUTCOME MEASURES: Changes in serum/plasma creatinine, BUN, urinary protein, and glucose excretion of sevoflurane at 24 and 72-hours were determined. RESULTS: Six relevant studies were qualified by both the inclusion criteria and inclusion dates. This review consists of 873 patients, 65% are males and 35% are females, with mean age of 56 ± 3 years. Sevoflurane was compared to isoflurane with regard to its nephrotoxic potential. Analyses on the effects of sevoflurane were performed on serum/plasma creatinine, BUN, urinary protein, and glucose excretion at 24 and 72 hours which showed no statistical difference between sevoflurane and isoflurane. CONCLUSION: In an apparently healthy adult without coexisting renal disorder, sevoflurane does not produce elevations in creatinine and BUN above the established upper limit of the reference range.

A comparison of renal responses to sevoflurane and isoflurane in patients undergoing donor nephrectomy: a randomized controlled trial.

Sevoflurane and isoflurane are volatile halogenated ether widely used in anesthesia. Both have comparable potency and easy titratability but sevoflurane has lower pungency and results in faster patient recovery. Isoflurane, however, is more affordable. The nephrotoxicity of sevoflurane is undisputed but studies on isoflurane nephrotoxicity are lacking. The objective of this paper is to determine the overall nephrotoxicity profile of sevoflurane and isoflurane in donor nephrectomy patients using the renal function markers - nuclear glomerular filtration rate (GFR), serum creatinine, urine protein-to-creatinine ratio, proteinuria, and glucosuria. A randomized comparative study of postoperative renal functions in donor nephrectomy patients who had received either low-flow (< 1 L/min) sevoflurane or isoflurane were analyzed. The renal parameters were repeated 72 hours post anesthesia. Forty-seven subjects (46%) were randomized to receive isoflurane while fifty-five received sevoflurane (54%). Between the two anesthetic groups, there was no significant difference in terms of serum creatinine, total GFR, or nuclear GFR. There was a statistically higher proportion of patients with urine protein-to-creatinine ratios of 0.2 and above in the isoflurane group (64% vs. 38%), while more patients in the sevoflurane group had ratios above 0.2 (62% vs. 36%, P < 0.05). The type of anesthetic agent was not an independent predictor of increasing serum creatinine, total GFR and urine protein-to-creatinine ratio and nuclear GFR. In conclusion, the overall nephrotoxicity profile of sevoflurane and isoflurane-treated donor nephrectomy patients is minimal.

Cox1 mutation abrogates need for Cox23 in cytochrome c oxidase biogenesis.

Cox23 is a known conserved assembly factor for cytochrome c oxidase, although its role in cytochrome c oxidase (CcO) biogenesis remains unresolved. To gain additional insights into its role, we isolated spontaneous suppressors of the respiratory growth defect in cox23∆ yeast cells. We recovered independent colonies that propagated on glycerol/lactate medium for cox23∆ cells at 37°C. We mapped these mutations to the mitochondrial genome and specifically to COX1 yielding an I101F substitution. The I101F Cox1 allele is a gain-of-function mutation enabling yeast to respire in the absence of Cox23. CcO subunit steady-state levels were restored with the I101F Cox1 suppressor mutation and oxygen consumption and CcO activity were likewise restored. Cells harboring the mitochondrial genome encoding I101F Cox1 were used to delete genes for other CcO assembly factors to test the specificity of the Cox1 mutation as a suppressor of cox23∆ cells. The Cox1 mutant allele fails to support respiratory growth in yeast lacking Cox17, Cox19, Coa1, Coa2, Cox14 or Shy1, demonstrating its specific suppressor activity for cox23∆ cells.

Disruption of a tight cluster surrounding tyrosine 131 in the native conformation of antithrombin III activates it for factor Xa inhibition.

The native conformation of antithrombin III (ATIII) is a poor inhibitor of its coagulation pathway target enzymes because of the partial insertion of its reactive center loop (RCL) in its central A beta-sheet. This study focused on tyrosine 131, which is located at the helix D-sheet A interface, adjacent to the ATIII pentasaccharide and heparin cofactor-binding sites and some 17A away from the RCL insertion. Crystallographic structures show that the Tyr(131) ring is buried in native ATIII and then becomes exposed when pentasaccharide binds to the inhibitor and activates it. This change suggested that Tyr(131) might serve as a switch for ATIII conformational activation. The hypothesis is supported by results from this study, which progressively removed atoms from the Tyr(131) side chain. Rates of heparin-independent Y131L and Y131A factor Xa inhibition were 25 and 29 times faster than for the control and Y131F, suggesting that Tyr(131) ring interactions with neighboring helix D and strand 2A residues shift the uncatalyzed native-to-activated conformational equilibrium toward the RCL-inserted state. Thermal denaturation experiments showed Y131A and Y131L were less stable than the control and Y131F, implying an increased tendency toward A-sheet mobility in these genetically activated molecules. Thus, the tight Tyr(131)-Asn(127)-Leu(130)-Leu(140)-Ser(142) cluster at the helix D-strand 2A interface of native antithrombin contributes significantly to the stability of the ground state conformation, and tyrosine 131 serves as a heparin-responsive molecular switch during the allosteric activation of ATIII anticoagulant activity.