Beta-propeller Protein Research Articles

BPAN manifesting with febrile seizures and language delay:a case report from Brazil

Neurodegeneration with brain iron accumulation (NBIA) is a complex group of hereditary progressive neurodegenerative diseases characterized by deposition of iron in the basal ganglia. Twelve genetic forms of this disorder have been identified in previous studies. Though they have different inheritance mechanisms all are usually associated with abnormal brain MRI findings. One of NBIA types is an X-linked disorder known as Beta-propeller Protein Associated Neurodegeneration (BPAN). Herein we describe the case of a 4-year-old girl with 2 episodes of febrile seizures, a brain MRI showing nonspecific hyperintense signal in the dentate nucleus area, and delays in language and communication development. Her diagnosis was made based on a genetic evaluation where exome sequencing revealed a mutation in the position chrX:48.933.022 region of the WDR45 gene. The literature describes different clinical presentations for BPAN, each with a different prognosis, suggesting a wide range of possible symptoms of BPAN, including mild cognitive delay and even epileptic encephalopathy (EE).

Neurodegeneration with brain iron accumulation: Characterization of clinical, radiological, and genetic features of pediatric patients from Southern India

BackgroundNeurodegeneration with brain iron accumulation (NBIA) is a group of rare inherited neurodegenerative disorders. Ten types of NBIA are known. Studies reporting various NBIA subtypes together are few. This study was aimed at describing clinical features, neuroimaging findings, and genetic mutations of different NBIA group disorders. MethodsClinical, radiological, and genetic data of patients diagnosed with NBIA in a tertiary care centre in Southern India from 2014 to 2020 was retrospectively collected and analysed. ResultsIn our cohort of 27 cases, PLA2G6-associated neurodegeneration (PLAN) was most common (n = 13) followed by Pantothenate kinase-associated neurodegeneration (PKAN) (n = 9). We had 2 cases each of Mitochondrial membrane-associated neurodegeneration (MPAN) and Beta-propeller protein- associated neurodegeneration (BPAN) and 1 case of Kufor-Rakeb Syndrome (KRS). Walking difficulty was the presenting complaint in all PKAN cases, whereas the presentation in PLAN was that of development regression with onset at a mean age of 2 years. Overall, 50% patients of them presented with development regression and one-third had epilepsy. Presence of pyramidal signs was most common examination feature (89%) followed by one or more eye findings (81%) and movement disorders (50%). Neuroimaging was abnormal in 24/27 cases and cerebellar atrophy was the commonest finding (52%) followed by globus pallidus hypointensities (44%). ConclusionsOne should have a high index of clinical suspicion for the diagnosis of NBIA in children presenting with neuroregression and vision abnormalities in presence of pyramidal signs or movement disorders. Neuroimaging and ophthalmological evaluation provide important clues to diagnosis in NBIA syndromes.

Clinical features and blood iron metabolism markers in children with beta-propeller protein associated neurodegeneration.

Neurodegeneration with brain iron accumulation constitutes a group of rare progressive movement disorders sharing intellectual disability and neuroimaging findings as common denominators. Beta-propeller protein-associated neurodegeneration (BPAN) represents approximately 7% of the cases, and its first signs are typically epilepsy and developmental delay. We aimed to describe in detail the phenotype of BPAN with a special focus on iron metabolism. We present a cohort of paediatric patients with pathogenic variants of WD-Repeat Domain 45 gene (WDR45). The diagnosis was established by targeted panel sequencing of genes associated with epileptic encephalopathies (n=9) or by Sanger sequencing of WDR45 (n=1). Data on clinical characteristics, molecular-genetic findings and other performed investigations were gathered from all participating centres. Markers of iron metabolism were analysed in 6 patients. Ten children (3 males, 7 females, median age 8.4 years) from five centres (Prague, Berlin, Vogtareuth, Tubingen and Cologne) were enrolled in the study. All patients manifested first symptoms (e.g. epilepsy, developmental delay) between 2 and 31 months (median 16 months). Seven patients were seizure-free (6 on antiepileptic medication, one drug-free) at the time of data collection. Neurological findings were non-specific with deep tendon hyperreflexia (n=4) and orofacial dystonia (n=3) being the most common. Soluble transferrin receptor/log ferritin ratio was elevated in 5/6 examined subjects; other parameters of iron metabolism were normal. Severity of epilepsy often gradually decreases in BPAN patients. Elevation of soluble transferrin receptor/log ferritin ratio could be another biochemical marker of the disease and should be explored by further studies.

Atg21 organizes Atg8 lipidation at the contact of the vacuole with the phagophore

ABSTRACT Coupling of Atg8 to phosphatidylethanolamine is crucial for the expansion of the crescent-shaped phagophore during cargo engulfment. Atg21, a PtdIns3P-binding beta-propeller protein, scaffolds Atg8 and its E3-like complex Atg12–Atg5-Atg16 during lipidation. The crystal structure of Atg21, in complex with the Atg16 coiled-coil domain, showed its binding at the bottom side of the Atg21 beta-propeller. Our structure allowed detailed analyses of the complex formation of Atg21 with Atg16 and uncovered the orientation of the Atg16 coiled-coil domain with respect to the membrane. We further found that Atg21 was restricted to the phagophore edge, near the vacuole, known as the vacuole isolation membrane contact site (VICS). We identified a specialized vacuolar subdomain at the VICS, typical of organellar contact sites, where the membrane protein Vph1 was excluded, while Vac8 was concentrated. Furthermore, Vac8 was required for VICS formation. Our results support a specialized organellar contact involved in controlling phagophore elongation. Abbreviations: FCCS: fluorescence cross correlation spectroscopy; NVJ: nucleus-vacuole junction; PAS: phagophore assembly site; PE: phosphatidylethanolamine;PROPPIN: beta-propeller that binds phosphoinositides; PtdIns3P: phosphatidylinositol-3-phosphate; VICS: vacuole isolation membrane contact site.

Serial MRI alterations of pediatric patients with beta-propeller protein associated neurodegeneration (BPAN)

Background and PurposeBeta-propeller protein-associated neurodegeneration (BPAN) is one subtype of neurodegeneration with brain iron accumulation. It is difficult to diagnose BPAN due to the non-specificity of their clinical findings and neuroimaging in early childhood. We experienced four pediatric patients with serial brain MRI and evaluated the alteration of the findings through their course. MethodsWe retrospectively reviewed the clinical findings and 21 MRI findings of the four patients with genetically confirmed pediatric BPAN. We also performed a quantitative MR assessment using the quantitative susceptibility mapping (QSM) values of the globus pallidus (GP), substantia nigra (SN), and deep cerebellar nuclei (DCN) compared to 10 age-matched disease controls. ResultsOnly one patient was suspected of BPAN based on imaging findings before the genetic diagnosis was made. The other three patients could not be suspected until their Whole-exome sequencings (WES) done. In all four cases, no abnormal signals were noted in the GP and SN at the initial brain MRI, but hypointensities were observed after the ages of 4–7 years on T2-weighted images and after the ages of 2–7 years on susceptibility-weighted images. In three patients, T2 hyperintensity in the bilateral DCN was persistently observed throughout the observational period. Three patients showed transient T2 hyperintensity and swelling in the GP, SN and/or DCN during the episodes of pyrexia and seizures. The other findings included cerebral and cerebellar atrophy, thinning of the corpus callosum, and delayed myelination. The QSM values of the GP and SN were significantly higher in the patients compared to the controls (P=0.005, respectively), but that of the DCN did not differ significantly (P=0.16). ConclusionBrain MRI is a useful method to establish the early diagnosis of BPAN.

Plant growth-promoting activity of beta-propeller protein YxaL secreted from Bacillus velezensis strain GH1-13.

YxaL is conserved within the Bacillus subtilis species complex associated with plants and soil. The mature YxaL protein contains a repeated beta-propeller domain, but the subcellular location and function of YxaL has not been determined. The gene encoding the mature YxaL protein was PCR amplified from genomic DNA of B. velezensis strain GH1-13 and used for recombinant protein production. A rabbit polyclonal antibody against the purified YxaL was generated and used for western blotting to determine the constitutive expression and secretion of YxaL. During normal culture growth of strain GH1-13, levels of the constitutively secreted YxaL were slowly rising to 100 μg L-1, and degraded with a half-life of 1.6 h in the culture medium. When the effects of YxaL on plant seed germination and seedling growth were examined, it was shown that seed treatment of Arabidopsis thaliana and rice (Oryza sativa L.) with purified YxaL at the optimal concentration of 1 mg L-1 was effective at improving the root growth of plants. Seedlings from the treated Arabidopsis seeds markedly increased transcription of a 1-aminocyclopropane-1-carboxylate synthetase marker gene (ACS11) but reduced expression of auxin- and abscisic acid-responsive marker genes (IAA1, GH3.3, and ABF4), especially when provided with exogenous auxin. Horticulture experiments showed that pepper (Capsicum annuum) seeds treated with 1 mg L-1 YxaL in a soaking solution increased shoot growth and improved tolerance to drought stress. We hypothesize that YxaL secreted from plant growth-promoting Bacillus cells has a significant impact on plant roots, with the potential to improve plant growth and stress tolerance.

A new complex rearrangement in infant ALL: t(X;11;17)(p11.2;q23;q12)

We present a case of an infant who developed pro-B acute lymphoblastic leukemia with a rare and complex MLL-translocation. Cytogenetic analysis of bone marrow cells at diagnosis showed a 46,XY,t(X;11)(p11.2;q23)[13]/46,XY[7] karyotype. Fluorescence in situ hybridization analysis using a break apart specific probes showed a split in the MLL gene. Long distance inverse-PCR and next generation sequencing analysis depicted a complex rearrangement t(X;11;17)(p11.2;q23;q12) involving MLL, MLLT6 and the genomic region Xp11.23, 41 bases upstream of the WDR45 gene. WDR45 encodes a beta-propeller protein essential for autophagocytosis. MLL rearrangements with involvement of Xp have not been previously described.

Ycf48 involved in the biogenesis of the oxygen-evolving photosystem II complex is a seven-bladed beta-propeller protein

Robust photosynthesis in chloroplasts and cyanobacteria requires the participation of accessory proteins to facilitate the assembly and maintenance of the photosynthetic apparatus located within the thylakoid membranes. The highly conserved Ycf48 protein acts early in the biogenesis of the oxygen-evolving photosystem II (PSII) complex by binding to newly synthesized precursor D1 subunit and by promoting efficient association with the D2 protein to form a PSII reaction center (PSII RC) assembly intermediate. Ycf48 is also required for efficient replacement of damaged D1 during the repair of PSII. However, the structural features underpinning Ycf48 function remain unclear. Here we show that Ycf48 proteins encoded by the thermophilic cyanobacterium Thermosynechococcus elongatus and the red alga Cyanidioschyzon merolae form seven-bladed beta-propellers with the 19-aa insertion characteristic of eukaryotic Ycf48 located at the junction of blades 3 and 4. Knowledge of these structures has allowed us to identify a conserved "Arg patch" on the surface of Ycf48 that is important for binding of Ycf48 to PSII RCs but also to larger complexes, including trimeric photosystem I (PSI). Reduced accumulation of chlorophyll in the absence of Ycf48 and the association of Ycf48 with PSI provide evidence of a more wide-ranging role for Ycf48 in the biogenesis of the photosynthetic apparatus than previously thought. Copurification of Ycf48 with the cyanobacterial YidC protein insertase supports the involvement of Ycf48 during the cotranslational insertion of chlorophyll-binding apopolypeptides into the membrane.

Clinical investigation of beta-propeller protein associated neurogeneration (BPAN)

Clinical investigation of beta-propeller protein associated neurogeneration (BPAN)

Ferrous Iron Up-regulation in Fibroblasts of Patients with Beta Propeller Protein-Associated Neurodegeneration (BPAN).

Mutations in WDR45 gene, coding for a beta-propeller protein, have been found in patients affected by Neurodegeneration with Brain Iron Accumulation, NBIA5 (also known as BPAN). BPAN is a movement disorder with Non Transferrin Bound Iron (NTBI) accumulation in the basal ganglia as common hallmark between NBIA classes (Hayflick et al., 2013). WDR45 has been predicted to have a role in autophagy, while the impairment of iron metabolism in the different NBIA subclasses has not currently been clarified. We found the up-regulation of the ferrous iron transporter (-)IRE/Divalent Metal Transporter1 and down-regulation of Transferrin receptor in the fibroblasts of two BPAN affected patients with splicing mutations 235+1G>A (BPAN1) and 517_519ΔVal 173 (BPAN2). The BPAN patients showed a concomitant increase of intracellular ferrous iron after starvation. An altered pattern of iron transporters with iron overload is highlighted in BPAN human fibroblasts, supporting for a role of DMT1 in NBIA. We here present a novel element, about iron accumulation, to the existing knowledge in field of NBIA. Attention is focused to a starvation-dependent iron overload, possibly accounting for iron accumulation in the basal ganglia. Further investigation could clarify iron regulation in BPAN.

Cartilage acidic protein 1, a new member of the beta-propeller protein family with amyloid propensity.

Cartilage acidic protein1 (CRTAC1) is an extracellular matrix protein of chondrogenic tissue in humans and its presence in bacteria indicate it is of ancient origin. Structural modeling of piscine CRTAC1 reveals it belongs to the large family of beta-propeller proteins that in mammals have been associated with diseases, including amyloid diseases such as Alzheimer's. In order to characterize the structure/function evolution of this new member of the beta-propeller family we exploited the unique characteristics of piscine duplicate genes Crtac1a and Crtac1b and compared their structural and biochemical modifications with human recombinant CRTAC1. We demonstrate that CRTAC1 has a beta-propeller structure that has been conserved during evolution and easily forms high molecular weight thermo-stable aggregates. We reveal for the first time the propensity of CRTAC1 to form amyloid-like structures, and hypothesize that the aggregating property of CRTAC1 may be related to its disease-association. We further contribute to the general understating of CRTAC1's and beta-propeller family evolution and function. Proteins 2017; 85:242-255. © 2016 Wiley Periodicals, Inc.

EPA-0225 – Neuropsychiatric phenotype of beta-propeller protein-associated neurodegeneration (BPAN)

Introduction Neurodegeneration with brain iron accumulation (NBIA) comprises an array of progressive brain disorders presenting with neurological and psychiatric symptoms, sometimes accompanied by intellectual impairment and behavioural problems. Clinically, according to time of onset and deterioration gradient, the NBIAs can be subdivided into two major groups: early onset / rapid, and late onset / slow. To date, nine genetic types of NBIA can be identified. Recently, a novel NBIA was described showing an additional specific neuroimaging pattern in the substantia nigra together with global cerebral atrophy, originally termed Static Encephalopathy of childhood with Neurodegeneration in Adulthood (SENDA). Shortly thereafter, it became clear that SENDA is caused by de novo heterozygous mutations in the WDR45-gene located in Xp11.2 that was named Beta-propeller Protein Associated Neurodegeneration (BPAN). Objectives Investigating the neuropsychiatric phenotype of BPAN. Aims Diagnostic evaluation of three adult female patients. Methods Detailed neuropsychiatric examination of 3 patients with genetically proven BPAN. Results All patients had a moderate to severe intellectual disability. In their early thirties autistic-like and depressive symptoms paralleled progressive motor and cognitive decline, ultimately resulting in wheelchair dependency. MRI of the brain demonstrated significant iron deposition in the globus pallidus and the substantia nigra as well as cerebral atrophy. Dystonia of the extremities and parkinsonian symptoms were most prominent. Treatment with levodopa/carbidopa was not effective. Conclusion The neuropsychiatric phenotype of BPAN is characterized by symptoms from the autistic and anxiety spectrum that, however, should be functionally interpreted as resulting form motor deterioration and severe cognitive inflexibility.

Beta-propeller protein-associated neurodegeneration (BPAN), a rare form of NBIA: Novel mutations and neuropsychiatric phenotype in three adult patients

Neurodegeneration with brain iron accumulation (NBIA) comprises a group of rare neuropsychiatric syndromes characterized by iron accumulation in the basal ganglia. The pantothenate kinase-associated neurodegeneration (PKAN) was the first NBIA form to be genetically identified almost 15 years ago. Nowadays, eight types can be genetically distinguished. More recently, a novel NBIA was delineated and termed Static Encephalopathy of childhood with Neurodegeneration in Adulthood (SENDA), characterized by early intellectual disability followed by delayed progressive motor and cognitive deterioration with an onset in the second to third decade. Very recently, mutations in the WD repeat-containing protein 45 (WDR45) gene located on Xp11.23 were shown to be the causal factor. The protein encoded by WDR45 propels protein interaction important for autophagy. This form was therefore retermed Beta-propeller Protein Associated Neurodegeneration (BPAN). Here, the first three Dutch patients with genetically proven BPAN are comprehensively described with respect to course and neurological as well as neuropsychiatric phenotypes. All three showed a characteristic delayed progression of neurological symptoms with parkinsonism and prominent dystonia. Treatment with levodopa/carbidopa had limited effects only. Neuropsychiatric symptoms within the autistic and affective spectrum were present in the early phase of the disease. The specific course and prognosis should implicate restrained psychopharmacological interventions.The clinical picture and imaging hallmarks are often highly suggestive and should lead to suspect this specific disorder. However, the identification of a WDR45 mutation is needed for a definite diagnosis of BPAN.

β-Propeller protein-associated neurodegeneration: a new X-linked dominant disorder with brain iron accumulation.

Neurodegenerative disorders with high iron in the basal ganglia encompass an expanding collection of single gene disorders collectively known as neurodegeneration with brain iron accumulation. These disorders can largely be distinguished from one another by their associated clinical and neuroimaging features. The aim of this study was to define the phenotype that is associated with mutations in WDR45, a new causative gene for neurodegeneration with brain iron accumulation located on the X chromosome. The study subjects consisted of WDR45 mutation-positive individuals identified after screening a large international cohort of patients with idiopathic neurodegeneration with brain iron accumulation. Their records were reviewed, including longitudinal clinical, laboratory and imaging data. Twenty-three mutation-positive subjects were identified (20 females). The natural history of their disease was remarkably uniform: global developmental delay in childhood and further regression in early adulthood with progressive dystonia, parkinsonism and dementia. Common early comorbidities included seizures, spasticity and disordered sleep. The symptoms of parkinsonism improved with l-DOPA; however, nearly all patients experienced early motor fluctuations that quickly progressed to disabling dyskinesias, warranting discontinuation of l-DOPA. Brain magnetic resonance imaging showed iron in the substantia nigra and globus pallidus, with a 'halo' of T1 hyperintense signal in the substantia nigra. All patients harboured de novo mutations in WDR45, encoding a beta-propeller protein postulated to play a role in autophagy. Beta-propeller protein-associated neurodegeneration, the only X-linked disorder of neurodegeneration with brain iron accumulation, is associated with de novo mutations in WDR45 and is recognizable by a unique combination of clinical, natural history and neuroimaging features.

Molecular Interfaces of the Galactose-binding Protein Tectonin Domains in Host-Pathogen Interaction

Beta-propeller proteins function in catalysis, protein-protein interaction, cell cycle regulation, and innate immunity. The galactose-binding protein (GBP) from the plasma of the horseshoe crab, Carcinoscorpius rotundicauda, is a beta-propeller protein that functions in antimicrobial defense. Studies have shown that upon binding to Gram-negative bacterial lipopolysaccharide (LPS), GBP interacts with C-reactive protein (CRP) to form a pathogen-recognition complex, which helps to eliminate invading microbes. However, the molecular basis of interactions between GBP and LPS and how it interplays with CRP remain largely unknown. By homology modeling, we showed that GBP contains six beta-propeller/Tectonin domains. Ligand docking indicated that Tectonin domains 6 to 1 likely contain the LPS binding sites. Protein-protein interaction studies demonstrated that Tectonin domain 4 interacts most strongly with CRP. Hydrogen-deuterium exchange mass spectrometry mapped distinct sites of GBP that interact with LPS and with CRP, consistent with in silico predictions. Furthermore, infection condition (lowered Ca(2+) level) increases GBP-CRP affinity by 1000-fold. Resupplementing the system with a physiological level of Ca(2+) did not reverse the protein-protein affinity to the basal state, suggesting that the infection-induced complex had undergone irreversible conformational change. We propose that GBP serves as a bridging molecule, participating in molecular interactions, GBP-LPS and GBP-CRP, to form a stable pathogen-recognition complex. The interaction interfaces in these two partners suggest that Tectonin domains can differentiate self/nonself, crucial to frontline defense against infection. In addition, GBP shares architectural and functional homologies to a human protein, hTectonin, suggesting its evolutionarily conservation for approximately 500 million years, from horseshoe crab to human.

Allosteric beta-propeller signalling in TolB and its manipulation by translocating colicins.

The Tol system is a five-protein assembly parasitized by colicins and bacteriophages that helps stabilize the Gram-negative outer membrane (OM). We show that allosteric signalling through the six-bladed beta-propeller protein TolB is central to Tol function in Escherichia coli and that this is subverted by colicins such as ColE9 to initiate their OM translocation. Protein-protein interactions with the TolB beta-propeller govern two conformational states that are adopted by the distal N-terminal 12 residues of TolB that bind TolA in the inner membrane. ColE9 promotes disorder of this 'TolA box' and recruitment of TolA. In contrast to ColE9, binding of the OM lipoprotein Pal to the same site induces conformational changes that sequester the TolA box to the TolB surface in which it exhibits little or no TolA binding. Our data suggest that Pal is an OFF switch for the Tol assembly, whereas colicins promote an ON state even though mimicking Pal. Comparison of the TolB mechanism to that of vertebrate guanine nucleotide exchange factor RCC1 suggests that allosteric signalling may be more prevalent in beta-propeller proteins than currently realized.

LGI1mutations in autosomal dominant and sporadic lateral temporal epilepsy

Autosomal dominant lateral temporal epilepsy (ADLTE) or autosomal dominant partial epilepsy with auditory features (ADPEAF) is an inherited epileptic syndrome with onset in childhood/adolescence and benign evolution. The hallmark of the syndrome consists of typical auditory auras or ictal aphasia in most affected family members. ADTLE/ADPEAF is associated in about half of the families with mutations of the leucine-rich, glioma-inactivated 1 (LGI1) gene. In addition, de novo LGI1 mutations are found in about 2% of sporadic cases with idiopathic partial epilepsy with auditory features, who are clinically similar to the majority of patients with ADLTE/ADPEAF but have no family history. Twenty-five LGI1 mutations have been described in familial and sporadic lateral temporal epilepsy patients. The mutations are distributed throughout the gene and are mostly missense mutations occurring in both the N-terminal leucine rich repeat (LRR) and C-terminal EPTP (beta propeller) protein domains. We show a tridimensional model of the LRR protein region that allows missense mutations of this region to be divided into two distinct groups: structural and functional mutations. Frameshift, nonsense and splice site point mutations have also been reported that result in protein truncation or internal deletion. The various types of mutations are associated with a rather homogeneous phenotype, and no obvious genotype-phenotype correlation can be identified. Both truncating and missense mutations appear to prevent secretion of mutant proteins, suggesting a loss of function effect of mutations. The function of LGI1 is unclear. Several molecular mechanisms possibly leading to lateral temporal epilepsy are illustrated and briefly discussed.

Analysis of active‐site amino‐acid residues of human serum paraoxonase using competitive substrates

Serum paraoxonase (PON1) is a calcium-dependent six-fold beta-propeller protein structurally similar to the di-isopropylfluorophosphatase (DFPase) found in the squid Loligo vulgaris. Human serum paraoxonase (HuPON1) has been shown to hydrolyze an array of substrates even though relatively little is known about its physiological role(s) or its catalytic mechanism. Through site-directed mutagenesis studies, designed from a DFPase-like homology model, and from a crystal structure of a hybrid PON1 molecule, amino-acid residues essential for enzyme function, including H115 and F222, have been identified. It was shown previously that, when H115 is replaced with tryptophan, the resulting enzyme hydrolyzes paraoxon but not phenyl acetate. This study shows that, when present simultaneously, phenyl acetate competitively inhibits paraoxon hydrolysis by H115W. Conversely, when F222 is replaced with tyrosine, mutant F222Y can hydrolyze phenyl acetate but not paraoxon. The presence of DFP, an inhibitor of both arylesterase and paraoxonase activities of wild-type HuPON1 (mean Ki=0.48+/-0.15 mM), has no effect on the ability of F222Y to catalyze the hydrolysis of phenyl acetate, suggesting that the F222Y mutant is unable to bind DFP. Together, the results suggest that, in wild-type HuPON1, H115 and F222 are important in determining substrate binding and specificity, but are not likely to be directly involved in substrate hydrolysis.

Modification of Cys-837 identifies an actin-binding site in the beta-propeller protein scruin.

In the acrosomal process of Limulus sperm, the beta-propeller protein scruin cross-links actin into a crystalline bundle. To confirm that scruin has the topology of a beta-propeller protein and to understand how scruin binds actin, we compared the solvent accessibility of cysteine residues in scruin and the acrosomal process by chemical modification with (1,5-IAEDANS). In soluble scruin, the two most reactive cysteines of soluble scruin are C837 and C900, whereas C146, C333, and C683 are moderately reactive. This pattern of reactivity is consistent with the topology of a typical beta-propeller protein; all of the reactive cysteines map to putative loops and turns whereas the unreactive cysteines lie within the predicted interior of the protein. The chemical reactivities of cysteine in the acrosomal process implicate C837 at an actin-binding site. In contrast to soluble scruin, in the acrosomal process, C837 is completely unreactive while the other cysteines become less reactive. Binding studies of chemically modified scruin correlate the extent of modification at C837 with the extent of inhibition of actin binding. Furthermore, peptides corresponding to residues flanking C837 bind actin and narrow a possible actin-binding region to a KQK sequence. On the basis of these studies, our results suggest that an actin-binding site lies in the C-terminal domain of scruin and involves a putative loop defined by C837.