Haspin is a nuclear and chromosomal-associated serine/threonine (S/T) kinase responsible for mitotic phosphorylation of histone H3 Thr-3. Haspin has recognizable similarities to eukaryotic protein kinase (ePK) folds, but its sequences are highly different, so its structural organization is very interesting. We report the 2.15-Å crystal structure of the human Haspin kinase domain. Haspin's ePK fold contains a series of insertions and deletions. This structure illustrates how Haspin escapes the classical activation scheme of most other kinases. The αC helix with a conserved glutamate is essential for catalysis, and it adopts its final active conformation within the kinase's leaflets. It is sandwiched between an alpha helix insert in front of the kinase domain and an activation fragment in an unprecedented conformation. Activation fragments without phosphorylation residues are stacked on the structurally abnormal αEF helix. Prominently protruding from the folded core, it forms a broad platform containing several residues related to substrate binding. Overall, the structure of the Haspin kinase domain reveals an active conformation that can be used for substrate recognition and phosphorylation in the absence of external regulators.
It has been reported that Haspin (haploid germ cell-specific nuclear protein kinase) is a serine / threonine kinase that may play a role in stopping and differentiating the cell cycle of haploid germ cells. In addition, Haspin mRNA can be detected in diploid cell lines and tissues. Here, Haspin-like proteins have been identified in several major eukaryotes, including yeast, plants, fruit flies, fish and mammals, and an expanded group of C. elegans. Haspin-like proteins have completed but divergent eukaryotic protein kinase domain sequences. Although clearly related to each other and to other eukaryotic protein kinases, Haspin-related proteins lack conservation of a subset of residues that are almost unchanged in known kinases and have unique insertion regions. In fact, phylogenetic analysis has shown that Haspin-like proteins form a new family of eukaryotic protein kinases different from previously defined. The identification of related proteins in model organisms provides some preliminary insights into their functional properties and will provide new experimental approaches for determining the function of Haspin proteins in mammalian cells. There are several atypical kinases in the human genome. Atypical kinases are homologous to the ePK family and may therefore adopt bi-leaf ePK folding, but they lack at least one conserved catalytic residue and may therefore be inactive by the enzyme, or their activation mechanism may be very different from typical kinases. Haspin/Gsg2 (haploid cell-specific nuclear protein kinase/germ cell-specific gene 2) is an atypical kinase with weak similarities to ePK profiles. Haspin family kinases have a similar domain structure. The kinase domain occupies the C-terminal portion of the molecule with low complexity and may have a partially unfolded N-terminal region, reducing sequence identity in the family. Haspin is a nuclear protein that is chromosomal-associated and preferentially phosphorylates histone H3 (P-Thr-3H3) threonine 3 within the mitochondria. Recently, this modification has been linked to the activation of Aurora B kinase, a key regulator of mitotic processes, but previous studies have failed to establish a convincing relationship between Haspin and Aurora B activity. Consistent with Haspin's mitotic effect, RNAi knockout will cause cell aggregation, chromosome cohesion and alignment problems, and metastasis.
The structure of Haspin's atypical kinase domain reveals an unprecedented activation mechanism. Even in the absence of activated ring phosphorylation, a wide range of hydrophobic interactions imparts a catalytically compatible conformation of the αC helix and the activating moiety, suggesting that Haspin is a constitutively active kinase. After removing at least four significant autophosphorylation sites, Haspin is active and therefore unlikely to be regulated by phosphorylation. The amino-terminal region of Haspin (1-450) is a low-complexity sequence and is unlikely to fold into a stable globular domain. Specific sequences in the Haspin N-terminal domain may regulate access to the substrate of the kinase domain in a manner similar to that recently described for the spindle checkpoint kinase Bub1, another atypical kinase.
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