Pcal_1699, an extremely thermostable malate dehydrogenase from hyperthermophilic archaeon Pyrobaculum calidifontis

Abstract

Two malate dehydrogenase homologs, Pcal_0564 and Pcal_1699, have been found in the genome of Pyrobaculum calidifontis. The gene encoding Pcal_1699 consisted of 927 nucleotides corresponding to a polypeptide of 309 amino acids. To examine the properties of Pcal_1699, the structural gene was cloned, expressed in Escherichia coli and the purified gene product was characterized. Pcal_1699 was NADH specific enzyme exhibiting a high malate dehydrogenase activity (886 U/mg) at optimal pH (10) and temperature (90 °C). Unfolding studies suggested that urea could not induce complete unfolding and inactivation of Pcal_1699 even at a final concentration of 8 M; however, in the presence of 4 M guanidine hydrochloride enzyme structure was unfolded with complete loss of enzyme activity. Thermostability experiments revealed that Pcal_1699 is the most thermostable malate dehydrogenase, reported to date, retaining more than 90 % residual activity even after heating for 6 h in boiling water.

Development and Implementation of Microbial Antifreeze Protein Based Coating for Anti‐Icing

“…[ 65,66 ] In addition, the insignificant number of hydrophobic residues such as Leucine (Leu), Valine (Val), and Isoleucine (Ile), which are particularly notable in highly stable proteins, is perhaps one of the main reasons for the low stability of this protein at higher temperatures. [ 67 ]…”

“…[65,66] In addition, the insignificant number of hydrophobic residues such as Leucine (Leu), Valine (Val), and Isoleucine (Ile), which are particularly notable in highly stable proteins, is perhaps one of the main reasons for the low stability of this protein at higher temperatures. [67] Repeating TxT sequences (T is Threonine and x is a nonconserved amino acid) have strong binding efficiency to the ice by anchored clathrate (AC) motifs. [68] The clathrate-like water molecules are formed around the methyl groups of the Thr residues by water molecules in the AC motifs and the bound hydroxyl group of the proteins.…”

Highly Active Thermophilic L-Asparaginase from Melioribacter roseus Represents a Novel Large Group of Type II Bacterial L-Asparaginases from Chlorobi-Ignavibacteriae-Bacteroidetes Clade

“…To adapt to elevated habitat temperatures, thermophilic L-asparaginases acquired a number of specific features, in particular, a low occurrence of thermolabile residues [42]. A correlation between thermostability and the ratio of preferred (Glu and Lys) and avoided (Gln and His) amino acids was reported [43,44].…”

Antifreeze Proteins: A Tale of Evolution From Origin to Energy Applications

“…Parameters such as temperature and pH change the protein stability and cause denaturation as a result of destabilizing bonds and altering charges. Thermostable proteins that are highly stable at 100 °C are reported to be predominant in hydrophobic residues, including valine leucine and isoleucine, and the higher number of disulfide bridges (Gharib et al, 2016;Gharib et al, 2020).…”