Ice nucleation (IN) active bacteria such as Pseudomonas syringae promote the growth of ice crystals more effectively than any material known. Using the specialized ice nucleation protein (INP) InaZ, P. syringae-the well studied epiphytic plant pathogen-attacks plants by frost damage and, likewise fascinating, drives ice nucleation within clouds when airborne in the atmosphere by linkage to the Earth's water cycle. While ice nucleation proteins play a tremendous role for life on the planet, the molecular details of their activity on the bacterial membrane surface are largely unknown. Bacterial ghosts (BGs) derived from Escherichia coli can be used as simplified model systems to study the mode of action of InaZ. In this work, the authors used BGs to study the role of InaZ localization on the luminal side of the bacterial inner membrane. Naturally, P. syringae INPs are displayed on the surface of the outer membrane; so in contrast, the authors engineered an N-terminal truncated form of inaZ lacking the transport sequence for anchoring of InaZ on the outer membrane. This construct was fused to N- and C-terminal inner membrane anchors and expressed in Escherichia coli C41. The IN activity of the corresponding living recombinant E. coli catalyzing interfacial ice formation of supercooled water at high subzero temperatures was tested by a droplet-freezing assay and surface spectroscopy. The median freezing temperature (T) of the parental living E. coli C41 cells without INP was detected at -20.1 °C and with inner membrane anchored INPs at a T value between -7 and -9 °C, demonstrating that the induction of IN from the inside of the bacterium by inner membrane anchored INPs facing the luminal inner membrane side is very similar to IN induced by bacterial INPs located at the outer membrane. Bacterial ghosts derived from these different constructs showed first droplet freezing values between -6 and -8 °C, whereas E. coli C41 BGs alone without carrying inner membrane anchored INPs exhibit a T of -18.9 °C. Sum frequency generation spectroscopy showed structural ordered water at the BG/water interface, which increased close to the water melting point. Together, this indicates that the more efficient IN of INP-BGs compared to their living parental strains can be explained by the free access of inner membrane anchored INP constructs to ultrapure water filling the inner space of the BGs.

Download full-text PDF

Source
http://dx.doi.org/10.1116/1.5142174DOI Listing

Publication Analysis

Top Keywords

inner membrane
32
ice nucleation
24
escherichia coli
16
membrane anchored
16
membrane
12
outer membrane
12
coli c41
12
anchored inps
12
inner
9
ice
8

Similar Publications

A rare haplotype of the GJD3 gene segregating in familial Meniere's disease interferes with connexin assembly.

Genome Med

January 2025

Otology & Neurotology Group CTS495, Instituto de Investigación Biosanitario, Ibs.GRANADA, Universidad de Granada, 18071, Granada, Spain.

Background: Familial Meniere's disease (FMD) is a rare polygenic disorder of the inner ear. Mutations in the connexin gene family, which encodes gap junction proteins, can also cause hearing loss, but their role in FMD is largely unknown.

Methods: We retrieved exome sequencing data from 94 individuals in 70 Meniere's disease (MD) families.

View Article and Find Full Text PDF

Cannabinoid receptor 1 (CB1R) has been extensively studied as a potential therapeutic target for various conditions, including pain management, obesity, emesis, and metabolic syndrome. Unlike orthosteric agonists such as Δ-tetrahydrocannabinol (THC), cannabidiol (CBD) has been identified as a negative allosteric modulator (NAM) of CB1R, among its other pharmacological targets. Previous computational and structural studies have proposed various binding sites for CB1R NAMs.

View Article and Find Full Text PDF

Chemistries on the inner leaflet of the cell membrane.

Chem Commun (Camb)

January 2025

Institute of Chemical Biology and Nanomedicine, State Key Laboratory of Chemo/Biosensing and Chemometrics, Hunan Provincial Key Laboratory of Biomacromolecular Chemical Biology, College of Chemistry and Chemical Engineering, Hunan University, Changsha, 410082, China.

The cell membrane, characterized by its inherent asymmetry, functions as a dynamic barrier that regulates numerous cellular activities. This Highlight aims to provide the chemistry community with a comprehensive overview of the intriguing and underexplored inner leaflet, encompassing both fundamental biology and emerging synthetic modification strategies. We begin by describing the asymmetric nature of the plasma membrane, with a focus on the distinct roles of lipids, proteins, and glycan chains, highlighting the composition and biofunctions of the inner leaflet and the biological mechanisms that sustain membrane asymmetry.

View Article and Find Full Text PDF

The permeability transition (PT) is a permeability increase of the mitochondrial inner membrane causing mitochondrial swelling in response to matrix Ca. The PT is mediated by regulated channel(s), the PT pore(s) (PTP), which can be generated by at least two components, adenine nucleotide translocator (ANT) and ATP synthase. Whether these provide independent permeation pathways remains to be established.

View Article and Find Full Text PDF

Rapid Preparation of Collagen/Red Blood Cell Membrane Tubes for Stenosis-Free Vascular Regeneration.

ACS Nano

January 2025

Shenzhen Key Laboratory of Smart Healthcare Engineering, Guangdong Provincial Key Laboratory of Advanced Biomaterials, Department of Biomedical Engineering, Southern University of Science and Technology, No. 1088 Xueyuan Road, Nanshan District, Shenzhen, Guangdong 518055, PR China.

Extracellular matrix (ECM)-based small-diameter vascular grafts (SDVGs, inner diameter (ID) < 6 mm) hold great promise for clinical applications. However, existing ECM-based SDVGs suffer from limited donor availability, complex purification, high cost, and insufficient mechanical properties. SDVGs with ECM-like structure and function, and good mechanical properties were rapidly prepared by optimizing common materials and preparation, which can improve their clinical prospects.

View Article and Find Full Text PDF

Want AI Summaries of new PubMed Abstracts delivered to your In-box?

Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!