Establishment of a Two-Dimensional HPLC-MS/MS Method Combined with DCl/D&lt;sub&gt;2&lt;/sub&gt;O Hydrolysis for the Determination of Trace Amounts of D-Amino Acid Residues in Proteins

Ishigo, Shoto; Negishi, E.; Miyoshi, Y.; Onigahara, Hirohisa; Mita, Masashi; Miyamoto, Tomofumi; Hara, Masaki; Homma, Hiroshi; Ueda, Tadashi; Hamase, Kenji

doi:10.15583/jpchrom.2015.017

Cited by 14 publications

(18 citation statements)

References 21 publications

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“…18,20−22 Using photolabile tags combined with electron capture dissociation MS, d -Ala, d -Ser, and d -Asp were identified in peptides of lens proteins. 18 Isomers of l -Asp, including d -Asp, l -iso-Asp, and d -iso-Asp, were distinguished in lens protein peptides using differential enzyme digestion with endoprotease Asp-N, protein- l -isoaspartyl methyltransferase, and paenidase ( d -aspartic acid endopeptidase).…”

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confidence: 99%

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A d-Amino Acid-Containing Neuropeptide Discovery Funnel

Livnat¹,

Tai²,

Jansson³

et al. 2016

Anal. Chem.

135

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A receptor binding class of d-amino acid-containing peptides (DAACPs) is formed in animals from an enzymatically mediated post-translational modification of ribosomally translated all-l-amino acid peptides. Although this modification can be required for biological actions, detecting it is challenging because DAACPs have the same mass as their all-l-amino acid counterparts. We developed a suite of mass spectrometry (MS) protocols for the nontargeted discovery of DAACPs and validated their effectiveness using neurons from Aplysia californica. The approach involves the following three steps, with each confirming and refining the hits found in the prior step. The first step is screening for peptides resistant to digestion by aminopeptidase M. The second verifies the presence of a chiral amino acid via acid hydrolysis in deuterium chloride, labeling with Marfey’s reagent, and liquid chromatography–mass spectrometry to determine the chirality of each amino acid. The third involves synthesizing the putative DAACPs and comparing them to the endogenous standards. Advantages of the method, the d-amino acid-containing neuropeptide discovery funnel, are that it is capable of detecting the d-form of any common chiral amino acid, and the first two steps do not require peptide standards. Using these protocols, we report that two peptides from the Aplysia achatin-like neuropeptide precursor exist as GdYFD and SdYADSKDEESNAALSDFA. Interestingly, GdYFD was bioactive in the Aplysia feeding and locomotor circuits but SdYADSKDEESNAALSDFA was not. The discovery funnel provides an effective means to characterize DAACPs in the nervous systems of animals in a nontargeted manner.

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confidence: 99%

“…21 Deuterium chloride-assisted acid hydrolysis was also used in conjunction with 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F) labeling to identify d -amino acid enantiomers of Ala, Asp, Glu, Pro, and Ser in peptides of ovalbumin. 22 …”

mentioning

confidence: 99%

A d-Amino Acid-Containing Neuropeptide Discovery Funnel

Livnat¹,

Tai²,

Jansson³

et al. 2016

Anal. Chem.

135

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show abstract

“…The Hamase group utilized a 2D‐LC strategy where d ‐amino acid residues in proteins were investigated by subjecting the sample to acid hydrolysis (cf. 5.2.1 Acid hydrolysis) followed by separation on a monolithic capillary octadecylsilica column and subsequent separation on Pirkle‐type narrowbore‐enantioselective columns, including KSAACSP‐001S and Sumichiral OA‐3200S as packing material . Using LC–MRM‐MS, they could distinguish five d / l isomers (Ala, Asp, Glu, Pro and Ser).…”

Section: Chromatographic Separationmentioning

confidence: 99%

“…If racemization occurs during acid hydrolysis in the presence of DCl/D 2 O, hydrogen–deuterium exchange takes place, which stably increases the mass of the amino acid by one mass unit (Fig. ) . Furthermore, because free amino acids are enantiomeric, the follow‐up analysis will require derivatization if an RP‐phase separation is desired.…”

Section: Determination Of Site‐specific Stereochemistry: Ms Tricksmentioning

confidence: 99%

Strategies for analysis of isomeric peptides

Jansson

2017

J of Separation Science

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This review presents an overview and recent progress of strategies for detecting isomerism in peptides, with focus on d/l epimerization and the various isomers that the presence of an aspartic acid residue may yield in a protein or peptide. While mass spectrometry has become a majorly used method of choice within proteomics, isomerism is inherently difficult to analyze because it is a modification that does not yield any change in mass of the analyte. Here, several techniques used for analysis of peptide isomerism are discussed, including enzymatic assays, liquid chromatography, and capillary electrophoresis. Recent progress in method development using mass spectrometry is also discussed, including labeling strategies, fragmentation techniques, and ion-mobility spectrometry.

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“…Concerning the protein-bound type D-enantiomers, D-amino acid residues are frequently observed in the "aged" proteins, and the correlation between D-amino acids and aging is a matter of high interest [21][22][23].…”

Section: Introductionmentioning

confidence: 99%

Sleep-Awake Profile Related Circadian D-Alanine Rhythm in Human Serum and Urine

et al. 2017

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The circadian profiles of D-alanine (Ala) amounts in human serum and urine have been clarified using a two-dimensional HPLC system combining a microbore reversed-phase C18 column and a Pirkle-type enantioselective column. The two-dimensional HPLC system enables the determination of trace amounts of D-Ala in the clinical samples without severe interference from intrinsic substances in the human physiological fluids. In the volunteers having normal sleep-awake patterns, the amounts of D-Ala in the serum were high in the late evening and low in the morning. On the other hand, in the volunteers having the reversed sleep-awake patterns, the amounts of D-Ala in the serum were high in the morning and low in the nighttime. The similar circadian changes of D-Ala amounts were observed in the urine. These 24-h profiles of D-Ala are almost the same as those observed in rats and mice, suggesting that D-Ala has fundamental physiological functions related to rest-active conditions in mammals, and could also be useful as the biomarker for sleep-awake profiles.

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Establishment of a Two-Dimensional HPLC-MS/MS Method Combined with DCl/D<sub>2</sub>O Hydrolysis for the Determination of Trace Amounts of D-Amino Acid Residues in Proteins

Cited by 14 publications

References 21 publications

A d-Amino Acid-Containing Neuropeptide Discovery Funnel

A d-Amino Acid-Containing Neuropeptide Discovery Funnel

Strategies for analysis of isomeric peptides

Sleep-Awake Profile Related Circadian D-Alanine Rhythm in Human Serum and Urine

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