Moore 2016 Cryobiology

From Bioblast
Publications in the MiPMap
Moore DS, Hand SC (2016) Cryopreservation of lipid bilayers by LEA proteins from Artemia franciscana and trehalose. Cryobiology 73:240-7.

Β» PMID: 27393243

Moore DS, Hand SC (2016) Cryobiology

Abstract: The capacity of Late Embryogenesis Abundant (LEA) proteins and trehalose to protect liposomes against freezing-induced damage was examined by measuring the leakage of 5(6)-carboxyfluorescein (CF). Liposomes were prepared to simulate the lipid compositions of the inner leaflet of the plasma membrane, outer mitochondrial membrane (OMM), and inner mitochondrial membrane (IMM). Two recombinant LEA proteins belonging to Group 3 (AfrLEA2 and AfrLEA3m) were expressed and purified from embryos of Artemia franciscana. Only OMM-like liposomes were significantly protected by AfrLEA2 and AfrLEA3m against freeze-thaw damage; at the highest protein:lipid mass ratio tested, leakage of CF was 56.3% of control with AfrLEA3m and 29.3% with AfrLEA2. By comparison, trehalose provided protection to all compositional types. The greatest stabilization during freezing occurred when trehalose was present on both sides of the bilayer. When mitochondria isolated from rat liver were freeze-thawed in trehalose solution, the OMM remained intact based on the absence of increased oxygen consumption when cytochrome c was added during oxidative phosphorylation (OXPHOS). Respiratory control ratios (OXPHOS/LEAK) were depressed by only 30% after freeze-thawing in trehalose compared to non-frozen controls, which indicated some retention of OXPHOS capacity by the IMM. Trehalose then was loaded into the matrix (0.24 ΞΌmol/mg mitochondrial protein) by transient opening of the permeability transition pore, a procedure optimized for retention of OMM integrity. Surprisingly, respiratory control ratios were not improved after freeze-thawing with external plus matrix trehalose, when compared to external trehalose alone. This result could perhaps be explained by insufficient accumulation of matrix trehalose.

Copyright Β© 2016 Elsevier Inc. All rights reserved. β€’ Keywords: Artemia franciscana, Freezing, Intrinsically disordered protein, Late embryogenesis abundant protein, Liposome, Mitochondria, Trehalose

β€’ O2k-Network Lab: US LA Baton Rouge Hand SC

Labels: MiParea: Respiration 

Stress:Cryopreservation  Organism: Rat  Tissue;cell: Liver  Preparation: Isolated mitochondria 

Coupling state: LEAK, OXPHOS, ET  Pathway:HRR: Oxygraph-2k 


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