Membranes Are Decisive for Maximum Freezing Efficiency of Bacterial Ice Nucleators.

R Schwidetzky,A T Backes,P Sudera,J Fröhlich-Nowoisky,K Meister,M Bonn,U Pöschl

doi:10.1021/acs.jpclett.1c03118

Abstract

Ice-nucleating proteins (INPs) from Pseudomonas syringae are among the most active ice nucleators known, enabling ice formation at temperatures close to the melting point of water. The working mechanisms of INPs remain elusive, but their ice nucleation activity has been proposed to depend on the ability to form large INP aggregates. Here, we provide experimental evidence that INPs alone are not sufficient to achieve maximum freezing efficiency and that intact membranes are critical. Ice nucleation measurements of phospholipids and lipopolysaccharides show that these membrane components are not part of the active nucleation site but rather enable INP assembly. Substantially improved ice nucleation by INP assemblies is observed for deuterated water, indicating stabilization of assemblies by the stronger hydrogen bonds of D2O. Together, these results show that the degree of order/disorder and the assembly size are critically important in determining the extent to which bacterial INPs can facilitate ice nucleation.

Highlights

Ice-nucleating proteins (INPs) from Pseudomonas syringae are among the most active ice nucleators known, enabling ice formation at temperatures close to the melting point of water
The INPs are localized in the outer bacterial membrane, as demonstrated by fractionating experiments and the isolation of ice-nucleating vesicles shed from the membrane.[8,14]
Recent studies further showed that environmental factors can have very different effects on class A and class C INs.[11,18−20] Here, we use the highthroughput twin-plate ice nucleation assay (TINA) to investigate the ice nucleation activity of the lipids 1,2dimyristoyl-3-trimethylammonium −propane (DPTAP), 1,2dipalmitoyl-sn-glycero-3-phosphoglycerol (DPPG), 1,2-dipalmitoyl-sn-glycero-3-phosphorylethanolamine (DPPE), phosphatidylinositol

Summary

Introduction

Ice-nucleating proteins (INPs) from Pseudomonas syringae are among the most active ice nucleators known, enabling ice formation at temperatures close to the melting point of water.

Results

Conclusion