The Role of Protein Loops and Linkers in Conformational Dynamics and Allostery

Papaleo, Elena; Saladino, Giorgio; Lambrughi, Matteo; Lindorff‐Larsen, Kresten; Gervasio, Francesco Luigi; Nussinov, Ruth

doi:10.1021/acs.chemrev.5b00623

Cited by 320 publications

(335 citation statements)

References 383 publications

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“…The transition between these two conformational states involves a rotation around helix ␣4 that closes the amino-terminal nucleotide binding domain. This hinge motion of helix ␣4 is similar to the rotation of the "␣C" helix in protein kinases (24). In the inactive conformation, the catalytic Glu-138 at the end of helix ␣4 is rotated out of the pocket, and in the active conformation, it rotates into the active site.…”

Section: Discussionmentioning

confidence: 75%

Allosteric Regulation of Mammalian Pantothenate Kinase

Subramanian

Yun²,

Yao

et al. 2016

Journal of Biological Chemistry

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Pantothenate kinase catalyzes the key regulatory step in CoA biosynthesis in bacteria and mammals (1-3). In mammals, there are four distinct kinases that exhibit tissue-specific expression as follows: PANK1␣ and PANK1␤ are splice variants of the PANK1 gene, and PANK2 and PANK3 are encoded by the PANK2 and PANK3 genes, respectively (4, 5). The kinase isoforms possess almost identical catalytic cores but differ in their amino termini that direct the enzymes to different subcellular compartments (6). Isoform 1␣ is targeted to the nucleus, whereas isoform 1␤ is associated with endosomes in humans and mice. Isoform 2 of mice is cytosolic, but the PANK2 gene in humans encodes a protein with both nuclear localization and mitochondrial targeting sequences. Isoform 3 is cytosolic in both species. All pantothenate kinases operate by a compulsory ordered mechanism with ATP⅐Mg 2ϩ as the leading substrate followed by pantothenate ( Fig. 1) (7). The principal mechanism for controlling mammalian pantothenate kinase activity is through feedback inhibition by acetyl-CoA, and the isoforms differ in their sensitivity to this regulator ( Fig. 1) (4, 8, 9). The 1␣ and 1␤ isoforms are least sensitive to inhibition, whereas isoforms 2 and 3 are more potently inhibited by acetyl-CoA. Acetyl-CoA inhibition is competitive with ATP⅐Mg 2ϩ , but acetyl-CoA binds far more tightly than ATP (10). Acyl-carnitines antagonize the inhibition of pantothenate kinases by acetyl-CoA (8).The importance of pantothenate kinase to mammalian physiology is highlighted by the phenotypes of knock-out mice and their connection to human disease. Isoform 1 is most highly expressed in the liver, and Pank1 Ϫ/Ϫ mice have lower total hepatic CoA and exhibit fatty acid ␤-oxidation and glucose homeostasis defects in the fasted state (11). In addition, Pank1 Ϫ/Ϫ Lep Ϫ/Ϫ mice have dramatically lower blood glucose and insulin levels compared with their diabetic Lep Ϫ/Ϫ counterparts, which highlights the connection between isoform 1 and glucose homeostasis (12). Consistent with this, an association between polymorphisms in the PANK1 gene and insulin levels was uncovered in a cohort of individuals in Finland (13). It has also been shown that the severe neurodegenerative disease pantothenate kinase-associated neurodegeneration arises from mutations in the human PANK2 gene (14). Unfortunately, Pank2 Ϫ/Ϫ mice do not recapitulate the pantothenate kinaseassociated neurodegeneration disease phenotype (15,16), and this may be due either to differences in the levels of isoform expression in mouse and human brains or to the fact that human PANK2 accumulates in the intra-membrane space in mitochondria, whereas mouse isoform 2 is cytosolic (9). Finally, Pank1 Ϫ/Ϫ Pank2 Ϫ/Ϫ double knock-out mice are unable to metabolize fats and ketones resulting in early postnatal death (16), and Pank1 Ϫ/Ϫ Pank3 Ϫ/Ϫ and Pank2 Ϫ/Ϫ Pank3 Ϫ/Ϫ double knock-out mice are both embryonic lethal. A chemical knockout of all pantothenate kinases in adult mice resulted in an 80% reduction in hepatic CoA levels a...

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Section: Discussionmentioning

confidence: 75%

Allosteric Regulation of Mammalian Pantothenate Kinase

Subramanian

Yun²,

Yao

et al. 2016

Journal of Biological Chemistry

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“…Having recognised its importance and conservation in the archaeal and human proteins, we explored whether the ‘wedge strategy’ was common to FEN1 enzymes in general, noting that the loop incorporating this residue meets the formal definition of an omega-loop (Supplementary Figure S6C–E): a protein structural element often associated with recognition processes (37,38). Sequence analysis confirmed the importance of this feature in FENs (Figure 6A and Supplementary Figure S11A) revealing strong conservation of both the wedge residue and C-terminal portion of the loop, which folds to form the core of the 3′-flap pocket upon binding.…”

Section: Resultsmentioning

confidence: 99%

A conserved loop–wedge motif moderates reaction site search and recognition by FEN1

Thompson

Gotham

Ciani

et al. 2018

Nucleic Acids Research

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DNA replication and repair frequently involve intermediate two-way junction structures with overhangs, or flaps, that must be promptly removed; a task performed by the essential enzyme flap endonuclease 1 (FEN1). We demonstrate a functional relationship between two intrinsically disordered regions of the FEN1 protein, which recognize opposing sides of the junction and order in response to the requisite substrate. Our results inform a model in which short-range translocation of FEN1 on DNA facilitates search for the annealed 3′-terminus of a primer strand, which is recognized by breaking the terminal base pair to generate a substrate with a single nucleotide 3′-flap. This recognition event allosterically signals hydrolytic removal of the 5′-flap through reaction in the opposing junction duplex, by controlling access of the scissile phosphate diester to the active site. The recognition process relies on a highly-conserved ‘wedge’ residue located on a mobile loop that orders to bind the newly-unpaired base. The unanticipated ‘loop–wedge’ mechanism exerts control over substrate selection, rate of reaction and reaction site precision, and shares features with other enzymes that recognize irregular DNA structures. These new findings reveal how FEN1 precisely couples 3′-flap verification to function.

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“…on loop motions (for review see 32). Here we have addressed the functional importance of the partially intrinsically disordered and highly conserved Tyr 138 ‐loop in the full‐length hPAH homotetramer 9, which was seen to have missing electron densities of the residues S 137 YGAEL in the unliganded rPAH tetramer 19.…”

Section: Discussionmentioning

confidence: 99%

Substituting Tyr¹³⁸ in the active site loop of human phenylalanine hydroxylase affects catalysis and substrate activation

et al. 2017

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Mammalian phenylalanine hydroxylase ( PAH ) is a key enzyme in l ‐phenylalanine ( l ‐Phe) metabolism and is active as a homotetramer. Biochemical and biophysical work has demonstrated that it cycles between two states with a variably low and a high activity, and that the substrate l ‐Phe is the key player in this transition. X‐ray structures of the catalytic domain have shown mobility of a partially intrinsically disordered Tyr 138 ‐loop to the active site in the presence of l ‐Phe. The mechanism by which the loop dynamics are coupled to substrate binding at the active site in tetrameric PAH is not fully understood. We have here conducted functional studies of four Tyr 138 point mutants. A high linear correlation ( r 2 = 0.99) was observed between their effects on the catalytic efficiency of the catalytic domain dimers and the corresponding effect on the catalytic efficiency of substrate‐activated full‐length tetramers. In the tetramers, a correlation ( r 2 = 0.96) was also observed between the increase in catalytic efficiency (activation) and the global conformational change (surface plasmon resonance signal response) at the same l ‐Phe concentration. The new data support a similar functional importance of the Tyr 138 ‐loop in the catalytic domain and the full‐length enzyme homotetramer.

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The Role of Protein Loops and Linkers in Conformational Dynamics and Allostery

Cited by 320 publications

References 383 publications

Allosteric Regulation of Mammalian Pantothenate Kinase

Allosteric Regulation of Mammalian Pantothenate Kinase

A conserved loop–wedge motif moderates reaction site search and recognition by FEN1

Substituting Tyr¹³⁸ in the active site loop of human phenylalanine hydroxylase affects catalysis and substrate activation

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The Role of Protein Loops and Linkers in Conformational Dynamics and Allostery

Cited by 320 publications

References 383 publications

Allosteric Regulation of Mammalian Pantothenate Kinase

Allosteric Regulation of Mammalian Pantothenate Kinase

A conserved loop–wedge motif moderates reaction site search and recognition by FEN1

Substituting Tyr138 in the active site loop of human phenylalanine hydroxylase affects catalysis and substrate activation

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Substituting Tyr¹³⁸ in the active site loop of human phenylalanine hydroxylase affects catalysis and substrate activation