Divergent Evolution of Eukaryotic CC- and A-Adding Enzymes

Synthesis of the CCA end of essential tRNAs is performed either by CCA-adding enzymes or as a collaboration between enzymes restricted to CC- and A-incorporation. While the occurrence of such tRNA nucleotidyltransferases with partial activities seemed to be restricted to Bacteria, the first example of such split CCA-adding activities was reported in Schizosaccharomyces pombe. Here, we demonstrate that the choanoflagellate Salpingoeca rosetta also carries CC- and A-adding enzymes. However, these enzymes have distinct evolutionary origins. Furthermore, the restricted activity of the eukaryotic CC-adding enzymes has evolved in a different way compared to their bacterial counterparts. Yet, the molecular basis is very similar, as highly conserved positions within a catalytically important flexible loop region are missing in the CC-adding enzymes. For both the CC-adding enzymes from S. rosetta as well as S. pombe, introduction of the loop elements from closely related enzymes with full activity was able to restore CCA-addition, corroborating the significance of this loop in the evolution of bacterial as well as eukaryotic tRNA nucleotidyltransferases. Our data demonstrate that partial CC- and A-adding activities in Bacteria and Eukaryotes are based on the same mechanistic principles but, surprisingly, originate from different evolutionary events.

Download Full-text

Expression and Solution NMR Study of Multi-site Phosphomimetic Mutant BCL-2 Protein

Protein and Peptide Letters ◽

10.2174/0929866526666190327121225 ◽

2019 ◽

Vol 26 (6) ◽

pp. 449-457

Author(s):

Ting Song ◽

Keke Cao ◽

Yu dan Fan ◽

Zhichao Zhang ◽

Zong W. Guo ◽

...

Keyword(s):

Structural Change ◽

Structural Biology ◽

Quantum Coherence ◽

Structural Changes ◽

Solution Nmr ◽

Flexible Loop ◽

Loop Region ◽

Loop Domain ◽

Nmr Study ◽

Binding Groove

Background: The significance of multi-site phosphorylation of BCL-2 protein in the flexible loop domain remains controversial, in part due to the lack of structural biology studies of phosphorylated BCL-2. Objective: The purpose of the study is to explore the phosphorylation induced structural changes of BCL-2 protein. Methods: We constructed a phosphomietic mutant BCL-2(62-206) (t69e, s70e and s87e) (EEEBCL- 2-EK (62-206)), in which the BH4 domain and the part of loop region was truncated (residues 2-61) to enable a backbone resonance assignment. The phosphorylation-induced structural change was visualized by overlapping a well dispersed 15N-1H heteronuclear single quantum coherence (HSQC) NMR spectroscopy between EEE-BCL-2-EK (62-206) and BCL-2. Results: The EEE-BCL-2-EK (62-206) protein reproduced the biochemical and cellular activity of the native phosphorylated BCL-2 (pBCL-2), which was distinct from non-phosphorylated BCL-2 (npBCL-2) protein. Some residues in BH3 binding groove occurred chemical shift in the EEEBCL- 2-EK (62-206) spectrum, indicating that the phosphorylation in the loop region induces a structural change of active site. Conclusion: The phosphorylation of BCL-2 induced structural change in BH3 binding groove.

Download Full-text

Identification of a flexible loop region (297–313) of urokinase-type plasminogen activator, which helps determine its catalytic activity.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)39603-0 ◽

1998 ◽

Vol 273 (2) ◽

pp. 1268

Author(s):

Ziyong Sun ◽

Yongping Jiang ◽

Zhong Ma ◽

Hui Wu ◽

Bei-Fang Liu ◽

...

Keyword(s):

Catalytic Activity ◽

Plasminogen Activator ◽

Flexible Loop ◽

Loop Region ◽

Type Plasminogen Activator ◽

Urokinase Type Plasminogen Activator

Download Full-text

The final steps of [FeFe]-hydrogenase maturation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1908121116 ◽

2019 ◽

Vol 116 (32) ◽

pp. 15802-15810 ◽

Cited By ~ 7

Author(s):

Oliver Lampret ◽

Julian Esselborn ◽

Rieke Haas ◽

Andreas Rutz ◽

Rosalind L. Booth ◽

...

Keyword(s):

Functional Integration ◽

Structural Features ◽

Site Directed Mutagenesis ◽

Structural Reorganization ◽

Biologically Inspired ◽

Hydrogenase Maturation ◽

Flexible Loop ◽

Loop Region ◽

Shed Light ◽

Secondary Ligand

The active site (H-cluster) of [FeFe]-hydrogenases is a blueprint for the design of a biologically inspired H2-producing catalyst. The maturation process describes the preassembly and uptake of the unique [2FeH] cluster into apo-hydrogenase, which is to date not fully understood. In this study, we targeted individual amino acids by site-directed mutagenesis in the [FeFe]-hydrogenase CpI of Clostridium pasteurianum to reveal the final steps of H-cluster maturation occurring within apo-hydrogenase. We identified putative key positions for cofactor uptake and the subsequent structural reorganization that stabilizes the [2FeH] cofactor in its functional coordination sphere. Our results suggest that functional integration of the negatively charged [2FeH] precursor requires the positive charges and individual structural features of the 2 basic residues of arginine 449 and lysine 358, which mark the entrance and terminus of the maturation channel, respectively. The results obtained for 5 glycine-to-histidine exchange variants within a flexible loop region provide compelling evidence that the glycine residues function as hinge positions in the refolding process, which closes the secondary ligand sphere of the [2FeH] cofactor and the maturation channel. The conserved structural motifs investigated here shed light on the interplay between the secondary ligand sphere and catalytic cofactor.

Download Full-text

Widespread nocturnality of living birds stemming from their common ancestor

BMC Evolutionary Biology ◽

10.1186/s12862-019-1508-y ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 1

Author(s):

Yonghua Wu

Keyword(s):

Positive Selection ◽

Common Ancestor ◽

Activity Patterns ◽

Bright Light ◽

Night Vision ◽

Diel Activity ◽

Divergent Evolution ◽

Nocturnal Activity ◽

Evolutionary Origins ◽

Diel Activity Patterns

Abstract Background Many living birds exhibit some nocturnal activity, but the genetic basis and evolutionary origins of their nocturnality remain unknown. Results Here, we used a molecular phyloecological approach to analyze the adaptive evolution of 33 phototransduction genes in diverse bird lineages. Our results suggest that functional enhancement of two night-vision genes, namely, GRK1 and SLC24A1, underlies the nocturnal adaption of living birds. Further analyses showed that the diel activity patterns of birds have remained relatively unchanged since their common ancestor, suggesting that the widespread nocturnal activity of many living birds may largely stem from their common ancestor rather than independent evolution. Despite this evolutionary conservation of diel activity patterns in birds, photoresponse recovery genes were found to be frequently subjected to positive selection in diverse bird lineages, suggesting that birds generally have evolved an increased capacity for motion detection. Moreover, we detected positive selection on both dim-light vision genes and bright-light vision genes in the class Aves, suggesting divergent evolution of the vision of birds from that of reptiles and that different bird lineages have evolved certain visual adaptions to their specific light conditions. Conclusions This study suggests that the widespread nocturnality of extant birds has a deep evolutionary origin tracing back to their common ancestor.

Download Full-text

Contribution of the flexible loop region to the function of staphylococcal enterotoxin B

Protein Engineering Design and Selection ◽

10.1093/protein/gzq006 ◽

2010 ◽

Vol 23 (5) ◽

pp. 415-421 ◽

Cited By ~ 4

Author(s):

Saeko Yanaka ◽

Motonori Kudou ◽

Yoshikazu Tanaka ◽

Takumi Sasaki ◽

Sumiyo Takemoto ◽

...

Keyword(s):

Staphylococcal Enterotoxin ◽

Staphylococcal Enterotoxin B ◽

Flexible Loop ◽

Loop Region ◽

Enterotoxin B

Download Full-text

Structural and Biophysical Analyses of Human N-Myc Downstream-Regulated Gene 3 (NDRG3) Protein

Biomolecules ◽

10.3390/biom10010090 ◽

2020 ◽

Vol 10 (1) ◽

pp. 90

Author(s):

Kyung Rok Kim ◽

Kyung A. Kim ◽

Joon Sung Park ◽

Jun Young Jang ◽

Yuri Choi ◽

...

Keyword(s):

Cancer Metabolism ◽

Underlying Mechanism ◽

Catalytic Triad ◽

Molecular Characteristics ◽

Flexible Loop ◽

Loop Region ◽

Hydrolase Fold ◽

Structural Differences ◽

And Migration ◽

Proliferation And Migration

The N-Myc downstream-regulated gene (NDRG) family belongs to the α/β-hydrolase fold and is known to exert various physiologic functions in cell proliferation, differentiation, and hypoxia-induced cancer metabolism. In particular, NDRG3 is closely related to proliferation and migration of prostate cancer cells, and recent studies reported its implication in lactate-triggered hypoxia responses or tumorigenesis. However, the underlying mechanism for the functions of NDRG3 remains unclear. Here, we report the crystal structure of human NDRG3 at 2.2 Å resolution, with six molecules in an asymmetric unit. While NDRG3 adopts the α/β-hydrolase fold, complete substitution of the canonical catalytic triad residues to non-reactive residues and steric hindrance around the pseudo-active site seem to disable the α/β-hydrolase activity. While NDRG3 shares a high similarity to NDRG2 in terms of amino acid sequence and structure, NDRG3 exhibited remarkable structural differences in a flexible loop corresponding to helix α6 of NDRG2 that is responsible for tumor suppression. Thus, this flexible loop region seems to play a distinct role in oncogenic progression induced by NDRG3. Collectively, our studies could provide structural and biophysical insights into the molecular characteristics of NDRG3.

Download Full-text

Elena Gracheva: Ion channels run hot and cold

The Journal of Cell Biology ◽

10.1083/jcb.2096pi ◽

2015 ◽

Vol 209 (6) ◽

pp. 778-779 ◽

Cited By ~ 1

Author(s):

Caitlin Sedwick

Keyword(s):

Ion Channels ◽

Molecular Basis ◽

Evolutionary Origins

Gracheva studies the molecular basis and evolutionary origins of hibernation.

Download Full-text

Contributions of a unique β-clamp to substrate recognition illuminates the molecular basis of exolysis in ferulic acid esterases

Biochemical Journal ◽

10.1042/bj20151153 ◽

2016 ◽

Vol 473 (7) ◽

pp. 839-849 ◽

Cited By ~ 8

Author(s):

Robert J. Gruninger ◽

Chris Cote ◽

Tim A. McAllister ◽

D. Wade Abbott

Keyword(s):

Ferulic Acid ◽

Molecular Basis ◽

Structural Model ◽

Substrate Recognition ◽

Hydroxycinnamic Acids ◽

Flexible Loop

Ferulic acid esterases (FAEs) hydrolyse bonds between hemicellulose and hydroxycinnamic acids. We characterize an FAE that utilizes a flexible loop to bind substrate and that is essential for catalysis. We propose a structural model for endolytic compared with exolytic FAE activity.

Download Full-text