Pseudokinases: Update on Their Functions and Evaluation as New Drug Targets

Byrne, Dominic P.; Foulkes, Daniel M; Eyers, Patrick A.

doi:10.4155/fmc-2016-0207

Cited by 71 publications

(70 citation statements)

References 184 publications

(233 reference statements)

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“…In contrast, among 50 human pseudokinases, 33 contain a residue other than Asp at the equivalent position. Therefore, missing the Asp residue is considered a hallmark of pseudokinases, even though some pseudokinases were discovered to possess significant kinase catalytic activity . Whether Asp can be replaced by any other residue has not been fully tested in any kinase.…”

Section: Discussionmentioning

confidence: 99%

“…Therefore, missing the Asp residue is considered a hallmark of pseudokinases, even though some pseudokinases were discovered to possess significant kinase catalytic activity. [35][36][37][38] Whether Asp can be replaced by any other residue has not been fully tested in any kinase. The screening of an Asp386 mutant library indicated that no other residue was able to replace Asp386 to allow significant kinase activity in Src (>0.2% of wild-type).…”

Section: Discussionmentioning

confidence: 99%

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Structural versatility that serves the function of the HRD motif in the catalytic loop of protein tyrosine kinase, Src

Cui

Sun

2018

Protein Science

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Site‐directed mutagenesis is a traditional approach for structure–function analysis of protein tyrosine kinases, and it requires the generation, expression, purification, and analysis of each mutant enzyme. In this study, we report a versatile high throughput bacterial screening system that can identify functional kinase mutants by immunological detection of tyrosine phosphorylation. Two key features of this screening system are noteworthy. First, instead of blotting bacterial colonies directly from Agar plates to nitrocellulose membrane, the colonies were cultured in 96‐well plates, and then spotted in duplicate onto the membrane with appropriate controls. This made the screening much more reliable compared with direct colony blotting transfer. A second feature is the parallel use of a protein tyrosine phosphatase (PTP)‐expressing host and a non‐PTP‐expressing host. Because high activity Src mutants are toxic to the host, the PTP system allowed the identification of Src mutants with high activity, while the non‐PTP system identified Src mutants with low activity. This approach was applied to Src mutant libraries randomized in the highly conserved HRD motif in the catalytic loop, and revealed that structurally diverse residues can replace the His and Arg residues, while the Asp residue is irreplaceable for catalytic activity.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Structural versatility that serves the function of the HRD motif in the catalytic loop of protein tyrosine kinase, Src

Cui

Sun

2018

Protein Science

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“…Similarly, the HER3 pseudokinase, although reported to have low catalytic activity, serves mostly as an allosteric activator for other members of the EGFR family of receptor kinases (Jura et al, 2009; Shi et al, 2010). These studies have highlighted the importance of pseudokinases in human biology and their diverse signaling functions make them attractive drug targets (Bailey et al, 2015; Byrne et al, 2017). …”

Section: Introductionmentioning

confidence: 99%

Protein AMPylation by an Evolutionarily Conserved Pseudokinase

Sreelatha

Yee

López

et al. 2018

Cell

161

224

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Summary Approximately 10% of human protein kinases are believed to be inactive and named pseudokinases because they lack residues required for catalysis. Here we show that the highly conserved pseudokinase selenoprotein-O (SelO) transfers AMP from ATP to Ser, Thr and Tyr residues on protein substrates (AMPylation), uncovering a previously unrecognized activity for a member of the protein kinase superfamily. The crystal structure of a SelO homolog reveals a protein kinase-like fold with ATP flipped in the active site, thus providing a structural basis for catalysis. SelO pseudokinases localize to the mitochondria and AMPylate proteins involved in redox homeostasis. Consequently, SelO activity is necessary for the proper cellular response to oxidative stress. Our results suggest that AMPylation may be a more widespread post translational modification than previously appreciated and that pseudokinases should be analyzed for alternative transferase activities.

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“…This work shows that the various speculations about the biological role(s) of human and amniote PANK4s are not due to PANK activity as previously believed. HsPANK4 may have an alternative function(s) and should be considered a pseudoenzyme …”

Section: Introductionmentioning

confidence: 99%

“…HsPANK4 may have an alternative function(s) and should be considered a pseudoenzyme. [21][22][23][24]…”

Section: Introductionmentioning

confidence: 99%

Human pantothenate kinase 4 is a pseudo‐pantothenate kinase

et al. 2019

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Pantothenate kinase generates 4′‐phosphopantothenate in the first and rate‐determining step of coenzyme A (CoA) biosynthesis. The human genome encodes three well‐characterized and nearly identical pantothenate kinases (PANK1‐3) plus a putative bifunctional protein (PANK4) with a predicted amino‐terminal pantothenate kinase domain fused to a carboxy‐terminal phosphatase domain. Structural and phylogenetic analyses show that all active, characterized PANKs contain the key catalytic residues Glu138 and Arg207 (HsPANK3 numbering). However, all amniote PANK4s, including human PANK4, encode Glu138Val and Arg207Trp substitutions which are predicted to inactivate kinase activity. Biochemical analysis corroborates bioinformatic predictions—human PANK4 lacks pantothenate kinase activity. Introducing Glu138Val and Arg207Trp substitutions to the human PANK3 and plant PANK4 abolished their robust pantothenate kinase activity. Introducing both catalytic residues back into human PANK4 restored kinase activity, but only to a low level. This result suggests that epistatic changes to the rest of the protein already reduced the kinase activity prior to mutation of the catalytic residues in the course of evolution. The PANK4 from frog, an anamniote living relative encoding the catalytically active residues, had only a low level of kinase activity, supporting the view that HsPANK4 had reduced kinase activity prior to the catalytic residue substitutions in amniotes. Together, our data show that human PANK4 is a pseudo‐pantothenate kinase—a catalytically deficient variant of the catalytically active PANK4 found in plants and fungi. The Glu138Val and Arg207Trp substitutions in amniotes (HsPANK3 numbering) completely deactivated the pantothenate kinase activity that had already been reduced by prior epistatic mutations.

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Pseudokinases: Update on Their Functions and Evaluation as New Drug Targets

Cited by 71 publications

References 184 publications

Structural versatility that serves the function of the HRD motif in the catalytic loop of protein tyrosine kinase, Src

Structural versatility that serves the function of the HRD motif in the catalytic loop of protein tyrosine kinase, Src

Protein AMPylation by an Evolutionarily Conserved Pseudokinase

Human pantothenate kinase 4 is a pseudo‐pantothenate kinase

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