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Light-Induced Covalent Buffer Adducts to Histidine in a Model Protein

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Abstract

Purpose

Light is known to induce histidine (His) oxidation and His-His crosslinking in proteins. The crosslinking is resulted from the nucleophilic attack of a His to a photooxidized His from another protein. The goal of this work is to understand if covalent buffer adducts on His residues can be generated by light through similar mechanisms in nucleophilic buffers such as Tris and His.

Methods

A model protein (DNase) was buffer exchanged into nucleophilic buffers before light exposure. Photogenerated products were characterized by tryptic peptide mapping with mass spectrometry (MS) analysis. Several buffer adductions on His residues were identified after light exposure. To understand the influencing factors of such reactions, the levels of adducts were measured for six nucleophilic buffers on all His residues in DNase.

Results

The levels of adducts were found to correlate with the solvent accessibility of the His residue. The levels of adducts also correlate with the structure of the nucleophile, especially the steric restrictions of the nucleophile. The levels of adducts can be higher than that of other His photoreaction products, including photooxidation and crosslinking.

Conclusions

In nucleophilic buffers, light can induce covalently-linked adducts to His residues.

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Abbreviations

1O2 :

Singlet oxygen

ACES:

N-(2-Acetamido)-2-Aminoethanesulfonic acid

Arg:

Arginine

Asn:

Asparagine

Asp:

Aspartic acid

Bis-tris :

1,3-Bis(Tris(Hydroxymethyl)Methylamino

Bis-tris-propane:

1,3-Bis(Tris(Hydroxymethyl)Methylamino)Propane

CaCl2 :

Calcium chloride

CF:

Cystic fibrosis

CHO:

Chinese hamster ovary

CID:

Collision-induced dissociation

Cys:

Cystenine

Da:

Daltons

DTT:

Dithiothreitol

Fc:

Fragment, crystallizable

FcRn:

Neonatal Fc receptor

Gly:

Glysine

HCl:

Hydrochloric acid

HEPES:

2-[4-(2-Hydroxyethyl)Piperazin-1-Yl]Ethanesulfonic acid

His:

Histidine

HPLC:

High performance liquid chromatography

ICH:

International conference on harmonisation of technical requirements for registration of pharmaceuticals for human use

IgG:

Immunoglobin G

Lys:

Lysine

MAb:

Monoclonal antibody

MES:

2-(N-Morpholino)Ethanesulfonic acid

Met:

Methionine

MOE:

Molecular operating environment

MOPSO:

3-Morpholino-2-Hydroxypropanesulfonic acid

MS:

Mass spectrometry

NaCl:

Sodium chloride

NaOH:

Sodium hydroxide

PETG:

Polyethylene terephthalate glycol

PS-20:

Polysorbate-20

RhDNase:

Recombinant human DNase

SASA:

Solvent accessible surface area

TFA:

Trifluoroacetic acid

TPA:

Tissue plasminogen activator

Tris:

(2-Amino-2-(Hydroxymethyl)-1,3-Propanediol

Trp:

Tryptophan

Tyr:

Tyrosine

UV:

Ultraviolet

Vis:

Visible

XIC:

Extracted ion chromatogram

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Acknowledgments and Disclosures

The authors would like to thank the insightful discussion with Taylor Zhang, Camellia Zamiri and Bingchuan Wei.

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Authors and Affiliations

  1. Protein Analytical Chemistry, Genentech, Inc., 1 DNA Way, South San Francisco, California, 94080, USA

    Ming Lei, Cynthia Quan & Yung-Hsiang Kao

  2. Late Stage Pharmaceutical Development, Genentech, Inc., 1 DNA Way, South San Francisco, California, 94080, USA

    Y. John Wang

  3. Department of Pharmaceutical Chemistry, University of Kansas, 2095 Constant Avenue, Lawrence, Kansas, 66047, USA

    Christian Schöneich

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  1. Ming Lei
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Correspondence to Ming Lei or Christian Schöneich.

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Lei, M., Quan, C., Wang, Y.J. et al. Light-Induced Covalent Buffer Adducts to Histidine in a Model Protein. Pharm Res 35, 67 (2018). https://doi.org/10.1007/s11095-017-2339-4

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