Analysis of D-amino acid-containing neuropeptides in mollusks and rodents

Livnat, Itamar

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https://hdl.handle.net/2142/97668 Copy

Description

Title

Analysis of D-amino acid-containing neuropeptides in mollusks and rodents

Author(s)

Livnat, Itamar

Issue Date

2017-04-03

Director of Research (if dissertation) or Advisor (if thesis)

Sweedler, Jonathan V.

Doctoral Committee Chair(s)

Sweedler, Jonathan V.

Committee Member(s)

• Gillette, Martha U.
• Raetzman, Lori T.
• Rhodes, Justin S.

Department of Study

Molecular & Integrative Physl

Discipline

Molecular & Integrative Physi

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• Neurochemistry
• D-amino acid-containing peptides
• Neuropeptides
• Mass spectrometry (MS)
• Liquid chromatography
• Aplysia californica
• Liquid chromatography–mass spectrometry (LC-MS)
• Liquid chromatography–mass spectrometry/mass spectrometry (LC-MS/MS)

Abstract

Modern mass spectrometry (MS)-based neuropeptidomics continues to reveal an unprecedented level of chemical complexity in the nervous system on the peptide level, with sometimes hundreds of peptides expressed by or secreted onto specific structures. These neuropeptides have evolved for animal behaviors and regulating neuronal circuits. Neuropeptides exert their responses through GPCRs in most cases, and in order to interact with the GPCR, their structure is important. Their structures are not solely defined by their peptide sequence, but may instead depend on the myriad post-translational modifications (PTMs) they undergo. Thus, the hunt for neuropeptides must include the discovery of the exact chemical forms of the physiologically active peptides as these are required to achieve their receptor binding capability, degradation and uptake pathways, and hence bioactivity. Life, on a molecular level, has a stereoselective preference for many biochemical processes. This is especially true for protein-derived neuropeptides, which are translated by ribosomes using exclusively L-amino acids. However, D-amino acids are present in animal peptides, first evidenced by dermorphin, an analgesic skin peptide in Phyllomedusa sauvagei. Dermorphin’s existence as a D-amino acid-containing peptide (DAACP) proved not to be a singularity, but instead revealed a novel post-translational modification: peptide isomerization. Roughly 40 endogenous animal DAACPs have been found in 3 evolutionarily distinct phyla as toxins, hormones, and importantly for this work, neuropeptides. Isomerization alters the three-dimensional shape of a peptide. This change in shape has been associated with a change in bioactivity of a peptide, with the DAACP being the sole bioactive form. However, isomerization does not change the molecular weight of a peptide, making it difficult to detect using MS. Thus, bioactive peptides are likely hiding in neuropeptidomics study under the assumption that these peptides, like the majority of life, are formed solely of L-amino acids. Understanding the physiological purpose of DAACPs requires their further discovery. An approach to characterize unknown DAACPs, the DAACP discovery funnel, is presented in this work. It aims to solve this problem through a series of adapted MS-based analytical techniques capable of identifying potential DAACPs and assaying their amino acids for their chiral state. The discovery funnel is used to discover new DAACPs in the Aplysia californica nervous system and is applied, as pioneering work, to nervous system structures in rodents.

Graduation Semester

2017-05

Type of Resource

text

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Copyright and License Information

Copyright 2016 Itamar Livnat

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