A wealth of genotype-specific proteoforms fine-tunes hemoglobin scavenging by haptoglobin

Sem Tamara, Vojtech Franc, Albert J R Heck

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

The serum haptoglobin protein (Hp) scavenges toxic hemoglobin (Hb) leaked into the bloodstream from erythrocytes. In humans, there are two frequently occurring allelic forms of Hp, resulting in three genotypes: Homozygous Hp 1-1 and Hp 2-2, and heterozygous Hp 2-1. The Hp genetic polymorphism has an intriguing effect on the quaternary structure of Hp. The simplest form, Hp 1-1, forms dimers consisting of two α1β units, connected by disulfide bridges. Hp 2-1 forms mixtures of linear (α1)2(α2)n-2(β)n oligomers (n > 1) while Hp 2-2 occurs in cyclic (α2)n(β)n oligomers (n > 2). Different Hp genotypes bind Hb with different affinities, with Hp 2-2 being the weakest binder. This behavior has a significant influence on Hp's antioxidant capacity, with potentially distinctive personalized clinical consequences. Although Hp has been studied extensively in the past, the finest molecular details of the observed differences in interactions between Hp and Hb are not yet fully understood. Here, we determined the full proteoform profiles and proteoform assemblies of all three most common genetic Hp variants. We combined several state-of-the-art analytical methods, including various forms of chromatography, mass photometry, and different tiers of mass spectrometry, to reveal how the tens to hundreds distinct proteoforms and their assemblies influence Hp's capacity for Hb binding. We extend the current knowledge by showing that Hb binding does not just depend on the donor's genotype, but is also affected by variations in Hp oligomerization, glycosylation, and proteolytic processing of the Hp α-chain.

Original languageEnglish
Pages (from-to)15554–15564
Number of pages11
JournalProceedings of the National Academy of Sciences of the United States of America
Volume117
Issue number27
DOIs
Publication statusPublished - 7 Jul 2020

Keywords

  • haptoglobin
  • genetic polymorphism
  • hybrid mass spectrometry
  • proteinglycosylation
  • proteolytic processing

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