Aldehyde dehydrogenases engage in many cellular functions, however their dysfunction resulting in accumulation of their substrates can be cytotoxic. ALDHs are responsible for the NAD(P)-dependent ...oxidation of aldehydes to carboxylic acids, participating in detoxification, biosynthesis, antioxidant and regulatory functions. Severe diseases, including alcohol intolerance, cancer, cardiovascular and neurological diseases, were linked to dysfunctional ALDH enzymes, relating back to key enzyme structure. An in-depth understanding of the ALDH structure-function relationship and mechanism of action is key to the understanding of associated diseases. Principal structural features 1) cofactor binding domain, 2) active site and 3) oligomerization mechanism proved critical in maintaining ALDH normal activity. Emerging research based on the combination of structural, functional and biophysical studies of bacterial and eukaryotic ALDHs contributed to the appreciation of diversity within the superfamily. Herewith, we discuss these studies and provide our interpretation for a global understanding of ALDH structure and its purpose–including correct function and role in disease. Our analysis provides a synopsis of a common structure-function relationship to bridge the gap between the highly studied human ALDHs and lesser so prokaryotic models.
The use of enzymes in biochemical processes is of interest due to their ability to work under mild conditions while attaining high reaction rates. A limitation in the use of enzymes such as ...oxidoreductases on a large scale lies with their requirement for costly cofactors,
e.g.
NAD
+
, in stoichiometric quantities. Cofactor regeneration mechanisms using bienzymatic recycling systems is an attractive way to increase productivity and efficiency. The thermophilic enzyme aldehyde dehydrogenase (ALDH
Tt
) was immobilized directly from
E. coli
cell lysate, containing the expressed enzyme, onto Ni
2+
activated Sepharose®. The system displayed a rate of conversion of approx. 63% NAD
+
with reuse achievable for up to 5 cycles and residual activity of the enzyme upon storage of 93% after 7 days.
l
-Lactate dehydrogenase was immobilized in a second reactor module downstream of ALDH
Tt
via
two different methods, electrochemical entrapment in poly(3,4-ethylenedioxypyrrole) (PEDOP) and covalent attachment on glyoxyl agarose. Both reactors allowed for up to 100% conversion of NADH, however LDH@agarose proved superior in terms of reuse and storage. LDH@agarose displayed no reduction in activity after 6 cycles of use and retained 98% activity following 56 days storage. A coupled reactor containing immobilized ALDH
Tt
-LDH was operated with the substrates hexanal, benzaldehyde, terephthalaldehyde and
p
-tolualdehyde. A particular advantage of the system is its ability to preferentially oxidise a single aldehyde group in substrates containing two aldehyde functional groups. The reactor demonstrated efficient cofactor regeneration under continual operation for up 24 h, with enhanced product yields.
An immobilized bi-enzymatic (ALDH
Tt
-LDH) flow reactor has been develeoped for the selective oxidation of aldehydes.
The Cation Diffusion Facilitators (CDFs) form a family of membrane-bound proteins capable of transporting zinc and other heavy metal ions. Involved in metal tolerance/resistance by efflux of ions, ...CDF proteins share a two-modular architecture consisting of a transmembrane domain (TMD) and C-terminal domain (CTD) that protrudes into the cytoplasm. Discovery of a Zn2+ and Cd2+ CDF transporter from a marine bacterium Maricaulis maris that does not possess the CTD questions current perceptions regarding this family of proteins. This article describes a new, CTD-lacking subfamily of CDFs and our current knowledge about this family of proteins in the view of these findings.
Piezoelectricity, the linear relationship between stress and induced electrical charge, has attracted recent interest due to its manifestation in biological molecules such as synthetic polypeptides ...or amino acid crystals, including gamma (γ) glycine. It has also been demonstrated in bone, collagen, elastin and the synthetic bone mineral hydroxyapatite. Piezoelectric coefficients exhibited by these biological materials are generally low, typically in the range of 0.1-10 pm V
, limiting technological applications. Guided by quantum mechanical calculations we have measured a high shear piezoelectricity (178 pm V
) in the amino acid crystal beta (β) glycine, which is of similar magnitude to barium titanate or lead zirconate titanate. Our calculations show that the high piezoelectric coefficients originate from an efficient packing of the molecules along certain crystallographic planes and directions. The highest predicted piezoelectric voltage constant for β-glycine crystals is 8 V mN
, which is an order of magnitude larger than the voltage generated by any currently used ceramic or polymer.
Aldehyde dehydrogenases (ALDH) form a superfamily of dimeric or tetrameric enzymes that catalyze the oxidation of a broad range of aldehydes into their corresponding carboxylic acids with the ...concomitant reduction of the cofactor NAD(P) into NAD(P)H. Despite their varied polypeptide chain length and oligomerisation states, ALDHs possess a conserved architecture of three domains: the catalytic domain, NAD(P)
binding domain, and the oligomerization domain. Here, we describe the structure and function of the ALDH from Thermus thermophilus (ALDH
) which exhibits non-canonical features of both dimeric and tetrameric ALDH and a previously uncharacterized C-terminal arm extension forming novel interactions with the N-terminus in the quaternary structure. This unusual tail also interacts closely with the substrate entry tunnel in each monomer providing further mechanistic detail for the recent discovery of tail-mediated activity regulation in ALDH. However, due to the novel distal extension of the tail of ALDH
and stabilizing termini-interactions, the current model of tail-mediated substrate access is not apparent in ALDH
. The discovery of such a long tail in a deeply and early branching phylum such as Deinococcus-Thermus indicates that ALDH
may be an ancestral or primordial metabolic model of study. This structure provides invaluable evidence of how metabolic regulation has evolved and provides a link to early enzyme regulatory adaptations.
A reversible temperature-dependent high-spin to low-spin transition with
T
1/2
= −60 °C has been observed in the resonance Raman spectra of the equilibrium reduced and photoreduced heme
a
3
of the ...thermophilic
ba
3
heme-copper oxidoreductase. The transition is based on the frequency shifts of the spin-state marker bands
ν
2
(C
b
C
b
) and
ν
10
(C
a
C
m
) and is attributed to the displacement of the heme iron along the heme normal as a consequence of the Fe-Np repulsion at temperature below −40 °C which will increase the ligand field strength forcing the pairing of d electrons into the lower energy orbitals.
A reversible temperature-dependent high- to low-spin transition with
T
1/2
= −60 °C has been observed in the resonance Raman spectra of the equilibrium reduced and photoreduced heme
a
3
of the thermophilic
ba
3
heme-copper oxidoreductase.
While drug‐eluting stents containing anti‐proliferative agents inhibit proliferation of smooth muscle cells (SMCs), they also delay the regrowth of the endothelial cells which can result in ...subsequent development of restenosis. Acidic extracellular environments promote cell anchorage and migration by inducing conformational change in integrins, the main cell adhesion proteins. This study addresses the feasibility of a citric acid (CA) functionalized nitinol stent for improving vascular biocompatibility, specifically enhancing endothelialization. CA functionalized nitinol vascular stents are compared to commercial bare metal (Zilver Flex) and paclitaxel eluting stents (Zilver PTX) in terms of re‐endothelialization. To study the effect of stent coatings, a stent conditioned media methodology was developed in an attempt to represent in vivo conditions. Overall, distinct advantages of the CA functionalized nitinol stent over commercial Zilver PTX DES and Zilver Flex BMS stents in terms of endothelial cell adhesion, migration, and proliferation are reported. These novel findings indicate the potential of a CA functionalized stent to serve as a bioactive and therapeutic surface for re‐endothelialization, perhaps in combination with a SMC proliferation inhibitor coating, to prevent restenosis.
By combining X-ray crystallography, electron paramagnetic resonance techniques and density functional theory-based modelling, we provide evidence for a direct coordination of the product analogue, ...phosphate, to the molybdenum active site of a sulfite dehydrogenase. This interaction is mimicking the still experimentally uncharacterized reaction intermediate proposed to arise during the catalytic cycle of this class of enzymes. This work opens new perspectives for further deciphering the reaction mechanism of this nearly ubiquitous class of oxidoreductases.
We provide structural and spectroscopic evidence for a molybdenum-phosphate adduct mimicking a proposed reaction intermediate in the active site of a prokaryotic sulfite oxidizing enzyme.
Aldehyde dehydrogenases (ALDH), found in all kingdoms of life, form a superfamily of enzymes that primarily catalyse the oxidation of aldehydes to form carboxylic acid products, while utilising the ...cofactor NAD(P)
. Some superfamily members can also act as esterases using
-nitrophenyl esters as substrates. The ALDH
from
was recombinantly expressed in
and purified to obtain high yields (approximately 15-20 mg/L) and purity utilising an efficient heat treatment step coupled with IMAC and gel filtration chromatography. The use of the heat treatment step proved critical, in its absence decreased yield of 40% was observed. Characterisation of the thermophilic ALDH
led to optimum enzymatic working conditions of 50 °C, and a pH of 8. ALDH
possesses dual enzymatic activity, with the ability to act as a dehydrogenase and an esterase. ALDH
possesses broad substrate specificity, displaying activity for a range of aldehydes, most notably hexanal and the synthetic dialdehyde, terephthalaldehyde. Interestingly,
-substituted benzaldehydes could be processed efficiently, but
-substitution resulted in no catalytic activity. Similarly, ALDH
displayed activity for two different esterase substrates,
-nitrophenyl acetate and
-nitrophenyl butyrate, but with activities of 22.9% and 8.9%, respectively, compared to the activity towards hexanal.
Controlling the electromechanical response of piezoelectric biological structures including tissues, peptides, and amino acids provides new applications for biocompatible, sustainable materials in ...electronics and medicine. Here, the piezoelectric effect is revealed in another class of biological materials, with robust longitudinal and shear piezoelectricity measured in single crystals of the transmembrane protein ba3 cytochrome c oxidase from Thermus thermophilus. The experimental findings from piezoresponse force microscopy are substantiated using a range of control measurements and molecular models. The observed longitudinal and shear piezoelectric responses of ≈2 and 8 pm V−1, respectively, are comparable to or exceed the performance of commonly used inorganic piezoelectric materials including quartz, aluminum nitride, and zinc oxide. This suggests that transmembrane proteins may provide, in addition to physiological energy transduction, technologically useful piezoelectric material derived entirely from nature. Membrane proteins could extend the range of rationally designed biopiezoelectric materials far beyond the minimalistic peptide motifs currently used in miniaturized energy harvesters, and the finding of robust piezoelectric response in a transmembrane protein also raises fundamental questions regarding the molecular evolution, activation, and role of regulatory proteins in the cellular nanomachinery, indicating that piezoelectricity might be important for fundamental physiological processes.
Piezoelectricity, the functional solid‐state property that converts force into voltage and vice versa, is demonstrated in a transmembrane protein, completing a “missing link” in the hierarchy of biological materials. The magnitudes of the piezoelectric coefficients are revealed through comprehensively benchmarked experiments and extensive molecular modeling. The results confirm transmembrane proteins as a promising source of sustainable, eco‐friendly, high‐performance, piezoelectric materials.
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