Exploring the bioisosterism of proline residue in melanostatin neuropeptide using heteroaromatic scaffolds

The HIF (hypoxia-inducible factor) plays a central regulatory role in oxygen homoeostasis. HIF proteins are regulated by three Fe(II)- and α-KG (α-ketoglutarate)-dependent prolyl hydroxylase enzymes [PHD (prolyl hydroxylase domain) isoenzymes 1–3 or PHD1, PHD2 and PHD3] and one asparaginyl hydroxylase [FIH (factor inhibiting HIF)]. The prolyl hydroxylases control the abundance of HIF through oxygen-dependent hydroxylation of specific proline residues in HIF proteins, triggering subsequent ubiquitination and proteasomal degradation. FIH inhibits the HIF transcription activation through asparagine hydroxylation. Understanding the precise roles and regulation of these four Fe(II)- and α-KG-dependent hydroxylases is of great importance. In the present paper, we report the biochemical characterization of the first HIF protein substrates that contain the CODDD (C-terminal oxygen-dependent degradation domain), the NODDD (N-terminal oxygen-dependent degradation domain) and the CAD (C-terminal transactivation domain). Using LC-MS/MS (liquid chromatography–tandem MS) detection, we show that all three PHD isoenzymes have a strong preference for hydroxylation of the CODDD proline residue over the NODDD proline residue and the preference is observed for both HIF1α and HIF2α protein substrates. In addition, steady-state kinetic analyses show differential substrate selectivity for HIF and α-KG in reference to the three PHD isoforms and FIH.

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Effect of proline residue on the hydrolysis of substrates by thermolysin

Biochemical and Biophysical Research Communications ◽

10.1016/0006-291x(69)90798-0 ◽

1969 ◽

Vol 34 (5) ◽

pp. 719-724 ◽

Cited By ~ 15

Author(s):

Hiroshi Matsubara ◽

Alan Singer ◽

Richard M. Sasaki

Keyword(s):

Proline Residue ◽

Hydrolysis Of

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Imidazolone formation from protonated tetrapeptides: Effects of replacing a glycine by an alanine or proline residue

International Journal of Mass Spectrometry ◽

10.1016/j.ijms.2018.10.002 ◽

2019 ◽

Vol 435 ◽

pp. 69-77 ◽

Cited By ~ 1

Author(s):

Justin Kai-Chi Lau ◽

K.H. Brian Lam ◽

Cheuk-Kuen Lai ◽

Alan C. Hopkinson ◽

K.W. Michael Siu

Keyword(s):

Proline Residue

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Characteristic features of specific binding of pentapeptide HFPGP labeled at the C-terminal proline residue to rat forebrain plasma membranes

Doklady Biochemistry and Biophysics ◽

10.1134/s1607672914030077 ◽

2014 ◽

Vol 456 (1) ◽

pp. 101-103 ◽

Cited By ~ 3

Author(s):

T. V. Vyunova ◽

L. A. Andreeva ◽

K. V. Shevchenko ◽

V. P. Shevchenko ◽

M. Yu. Bobrov ◽

...

Keyword(s):

Plasma Membranes ◽

Specific Binding ◽

Proline Residue ◽

Rat Forebrain ◽

Characteristic Features

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The role of prolines and glycine in the transmembrane domain of LAT

10.1101/2020.08.10.244251 ◽

2020 ◽

Cited By ~ 1

Author(s):

Daniela Glatzová ◽

Harsha Mavila ◽

Maria Chiara Saija ◽

Tomáš Chum ◽

Lukasz Cwiklik ◽

...

Keyword(s):

Amino Acids ◽

Plasma Membrane ◽

Transmembrane Domain ◽

Transmembrane Helix ◽

Helical Structure ◽

Surface Expression ◽

Transmembrane Segment ◽

Proline Residue ◽

And Function ◽

The Impact

ABSTRACTLAT is a critical regulator of T cell development and function. It organises signalling events at the plasma membrane. However, the mechanism, which controls LAT localisation at the plasma membrane is not fully understood. Here, we studied the impact of helix-breaking amino acids, two prolines and one glycine, in the transmembrane segment on localisation and function of LAT. Using in silico analysis, confocal and superresolution imaging and flow cytometry we demonstrate that central proline residue destabilises transmembrane helix by inducing a kink. The helical structure and dynamics is further regulated by glycine and another proline residue in the luminal part of LAT transmembrane domain. Replacement of these residues with aliphatic amino acids reduces LAT dependence on palmitoylation for sorting to the plasma membrane. However, surface expression of these mutants is not sufficient to recover function of non-palmitoylated LAT in stimulated T cells. These data indicate that geometry and dynamics of LAT transmembrane segment regulate its localisation and function in immune cells.

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What Hinges on the S3 Hinge?: Effects of a Conserved Proline Residue on Gating Current in S4-Based Voltage Sensors

Biophysical Journal ◽

10.1016/j.bpj.2012.11.718 ◽

2013 ◽

Vol 104 (2) ◽

pp. 125a

Author(s):

Michael F. Priest ◽

Francisco Bezanilla

Keyword(s):

Gating Current ◽

Proline Residue ◽

Voltage Sensors

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Requirement of the C-terminal proline residue for stability of the Ca2+-activated photoprotein aequorin

Biochemical Journal ◽

10.1042/bj2930181 ◽

1993 ◽

Vol 293 (1) ◽

pp. 181-185 ◽

Cited By ~ 40

Author(s):

N J Watkins ◽

A K Campbell

Keyword(s):

Light Emission ◽

Specific Activity ◽

In Vitro Transcription ◽

Wild Type ◽

Proline Residue ◽

Long Term Stability ◽

Recombinant Aequorin

cDNA coding for the Ca(2+)-activated photoprotein aequorin from the jellyfish Aequorea victoria has been engineered to investigate the role of the C-terminal proline residue in bioluminescence. Recombinant aequorin proteins were synthesized by PCR followed by in vitro transcription/translation, and characterized by specific activity, stability, and affinity for coelenterazine. The C-terminal proline residue of aequorin was shown to be essential for the long-term stability of the bound coelenterazine. Aequorin minus proline had only 1% of the specific activity of the wild-type after 2 h, and was virtually inactive after 18 h. The instability of this variant was further demonstrated by re-activating with a coelenterazine analogue (epsilon-coelenterazine), where maximum reactivation was reached in 15 min, and the luminescent activity was almost completely abolished within 3 h. Replacement of the C-terminal proline residue with histidine or glutamic acid decreased the specific activity to 10 and 19% of that of the wild-type respectively. However these variants were also unstable, having t1/2 values of 2.4 h and 2.3 h respectively. Enhancement of the Ca(2+)-independent light emission when proline was replaced by histidine confirmed the stabilizing role of the C-terminal proline. No significant effect of removal of the C-terminal proline was detected on the affinity for coelenterazine.

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