Change in structure of the N‐terminal region of transthyretin produces change in affinity of transthyretin to T4 and T3

Prapunpoj, Porntip; Leelawatwatana, Ladda; Schreiber, Gerhard; Richardson, Samantha J.

doi:10.1111/j.1742-4658.2006.05404.x

Cited by 31 publications

(35 citation statements)

References 37 publications

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“…This segment is shorter and more hydrophilic in eutherians becoming longer and more hydrophobic in marsupials (with two additional amino acids), and in birds, reptile and amphibian (with three additional amino acids). These changes could be explained by splice site shifts of intron 1 as previously proposed ( So far, binding studies involving recombinant chimeric xenopus/crocodile TTR and recently with human/crocodile TTR provided experimental evidence that this region influences TH binding affinity of TTR (Prapunpoj et al, 2006, Prapunpoj et al, 2002.…”

Section: Role Of the N-terminal Region On Sbttr Bindingmentioning

confidence: 86%

Hormone affinity and fibril formation of piscine transthyretin: The role of the N-terminal

Morgado

Melo

Lundberg

et al. 2008

Molecular and Cellular Endocrinology

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SummaryTransthyretin (TTR) transports thyroid hormones (THs), thyroxine (T 4 ) and triiodothyronine (T 3 ) in the blood of vertebrates. TH-binding sites are highly conserved in vertebrate TTR however, piscine TTR has a longer N-terminus which is thought to influence TH-binding affinity and may influence TTR stability. We produced recombinant wild-type sea bream TTR (sbTTRWT) plus two mutants in which six (sbTTRM6) and twelve (sbTTRM12) N-terminal residues were removed. Ligandbinding studies revealed similar affinities for T 3 (Kd=10.6±1.7nM) and T 4 (Kd=9.8±0.97nM) binding to sbTTRWT. Affinity for THs was unaltered in sbTTRM12 but sbTTRM6 had poorer affinity for T 4 (Kd=252.3±15.8nM) implying that some residues in the N-terminus can influence T 4 binding. sbTTRM6 inhibited acid-mediated fibril formation in vitro as shown by fluorometric measurements using thioflavine-T. In contrast, fibril formation by sbTTRM12 was significant, probably due to decreased stability of the tetramer. Such studies also suggested that sbTTRWT is more resistant to fibril formation than human TTR.

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Section: Role Of the N-terminal Region On Sbttr Bindingmentioning

confidence: 86%

Hormone affinity and fibril formation of piscine transthyretin: The role of the N-terminal

Morgado

Melo

Lundberg

et al. 2008

Molecular and Cellular Endocrinology

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“…The sizes of the TTR subunits were obtained by comparing their relative mobilities with protein markers: phosphorylase b (94 kDa), bovine serum albumin (67 kDa), ovalbumin (43 kDa), carbonic anhydrase (30 kDa), soya bean trypsin inhibitor (20.1 kDa), and α-lactalbumin (14.4 kDa). Crossed-reactivity of the recombinant TTRs was determined by western www.scienceasia.org blotting, followed by enhanced chemiluminescence (ECL) detection as previously described 6,7 . The primary and secondary antibodies used were anticrocTTR antibody raised in rabbit (dilution 1:2500) and horseradish peroxidase-conjugated anti-rabbit IgG antibody (dilution 1:10 000), respectively.…”

Section: Analysis Of the Physicochemical Properties Of Ttrsmentioning

confidence: 99%

“…The induction of Pichia clones to produce all the studied recombinant and chimeric TTRs was performed as described previously 6,7 . In brief, the Pichia clones of crocTTR 17 , xenoN/crocTTR 17 and the two new chimeric crocTTRs were induced with 0.5% methanol for 3 days at 28°C in buffered complex glycerol or methanol medium (BMGY/BMMY).…”

Section: Production and Purification Of Recombinant Ttrsmentioning

confidence: 99%

“…The nucleotide sequences of the cDNAs were confirmed by sequencing before they were ligated into a Pichia expression vector, pPIC9. The recombinant vectors were transformed into P. pastoris and the transformant clones were selected as described previously 6,7 .…”

Section: Construction Cloning and Expression Of Ttr Cdnasmentioning

confidence: 99%

“…In addition, variations in the length of the C-terminal region of TTRs from pig 3 and microbats 4,5 compared with that from human have also been detected. The changes in length and hydrophobicity were demonstrated to affect the binding affinities of TTR for THs and retinol binding protein (RBP) 6,7 . Recently, the proteolytic activity of TTR was discovered and its participation in the biology of the nervous system and high density lipoprotein (HDL) was demonstrated 8,9 .…”

Section: Introductionmentioning

confidence: 99%

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Proteolytic activity of Crocodylus porosus transthyretin protease and role of the terminal polypeptide sequences

Leelawatwattana¹,

Praphanphoj²,

Prapunpoj³

2016

ScienceAsia

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Human transthyretin (TTR), a recently identified protease, participates in the biology of high density lipoprotein and in the nervous system. In the present study, we determined whether TTR from a non-mammal Crocodylus porosus (crocTTR) has proteolytic activity and whether the N-and C-termini of the TTR polypeptide affect the proteolytic activity. The proteolytic activity of crocTTR and three chimeric crocTTRs: xenoN/crocTTR (crocTTR in which the N-terminal sequence was replaced with that of Xenopus laevis TTR), pigC/crocTTR (crocTTR in which the C-terminal sequence was replaced with that of Sus scrofa TTR), and xenoN/pigC/crocTTR (crocTTR in which the N-and C-terminal sequences were replaced with that of X. laevis TTR and S. scrofa TTR, respectively) were studied and compared. Using either casein or apoAI as a substrate, crocTTR had a lower proteolytic activity than human TTR. Replacing the C-terminal sequence of crocTTR increased the activity (casein: 1008 ± 36 pmol/min; apoAI: 231 ± 43 pmol/min), whereas replacing the N-terminal sequence decreased the activity (casein: 299 ± 26 pmol/min; apoAI: 31.5 ± 2.0 pmol/min). The activity of xenoN/pigC/crocTTR (casein: 502 ± 11 pmol/min; apoAI: 371 ± 23 pmol/min) was higher than those of crocTTR or xenoN/crocTTR, but similar to that of pigC/crocTTR. These results are the first to show the proteolytic activity of reptile TTR, and that the activity is changed when the N-and/or C-terminal amino acid sequences of the TTR subunit are changed.

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Increasing the length and hydrophobicity of the C‐terminal sequence of transthyretin strengthens its binding affinity to retinol binding protein

et al. 2017

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Transthyretin ( TTR ) is a transporter for thyroid hormone ( TH ) and retinol, the latter via binding with retinol binding protein ( RBP ). Both the N‐terminal and C‐terminal regions of the TTR subunit are located in close proximity to the central binding channel for ligands. During the evolution of vertebrates, these regions changed in length and hydropathy. The changes in the N‐terminal sequence were demonstrated to affect the binding affinities for TH s and RBP . Here, the effects of changes in the C‐terminal sequence were determined. Three chimeric TTR s, namely pigC/hu TTR (human TTR with the C‐terminal sequence changed to that of Sus scrofa TTR ), xenoN/pigC/hu TTR (human TTR with the N‐terminal and C‐terminal sequences changed to those of Xenopus laevis and S. scrofa , respectively), and pigC/croc TTR ( Crocodylus porosus TTR with the C‐terminal sequence changed to that of S. scrofa TTR ), were constructed and their binding affinities for human RBP were determined at low TTR / RBP molar ratio using chemiluminescence immunoblotting. The binding dissociation constant ( K d ) values of pigC/hu TTR , xenoN/pigC/hu TTR and pigC/croc TTR were 3.20 ± 0.35, 1.53 ± 0.38 and 0.31 ± 0.04 μ m , respectively, and the K d values of human and C. porosus TTR were 4.92 ± 0.68 and 1.42 ± 0.45 μ m , respectively. These results demonstrate chimeric TTR s bound RBP with a higher strength than wild‐type TTR s, and the changes in the C‐terminal sequence of TTR had a positive effect on its binding affinity for RBP . In addition, changes to the N‐terminal and C‐terminal sequences showed comparable effects on the binding affinity.

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Change in structure of the N‐terminal region of transthyretin produces change in affinity of transthyretin to T4 and T3

Cited by 31 publications

References 37 publications

Hormone affinity and fibril formation of piscine transthyretin: The role of the N-terminal

Hormone affinity and fibril formation of piscine transthyretin: The role of the N-terminal

Proteolytic activity of Crocodylus porosus transthyretin protease and role of the terminal polypeptide sequences

Increasing the length and hydrophobicity of the C‐terminal sequence of transthyretin strengthens its binding affinity to retinol binding protein

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