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J. Biol. Chem., Vol. 279, Issue 43, 45219-45225, October 22, 2004

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Characteristic Recognition of N-Acetylgalactosamine by an Invertebrate C-type Lectin, CEL-I, Revealed by X-ray Crystallographic Analysis^*

Hajime Sugawara, Masami Kusunoki, Genji Kurisu¶, Tokiko Fujimoto||, Haruhiko Aoyagi||, and Tomomitsu Hatakeyama||**

From the Research Center for Structural and Functional Proteomics, Institute for Protein Research, Osaka University, 3-2 Yamada-oka, Suita, Osaka 565-0871, Japan, the Laboratory for Communication Mechanisms, RIKEN Plant Science Center, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan, and the ^||Department of Applied Chemistry, Faculty of Engineering, Nagasaki University, 1-14 Bunkyo-machi, Nagasaki 852-8521, Japan

Received for publication, August 3, 2004 , and in revised form, August 19, 2004.

	ABSTRACT

TOP ABSTRACT INTRODUCTION EXPERIMENTAL PROCEDURES RESULTS AND DISCUSSION REFERENCES

 
CEL-I is a C-type lectin, purified from the sea cucumber Cucumaria echinata, that shows a high specificity for N-acetylgalactosamine (GalNAc). We determined the crystal structures of CEL-I and its complex with GalNAc at 2.0 and 1.7 Å resolution, respectively. CEL-I forms a disulfide-linked homodimer and contains two intramolecular disulfide bonds, although it lacks one intramolecular disulfide bond that is widely conserved among various C-type carbohydrate recognition domains (CRDs). Although the sequence similarity of CEL-I with other C-type CRDs is low, the overall folding of CEL-I was quite similar to those of other C-type CRDs. The structure of the complex with GalNAc revealed that the basic recognition mode of GalNAc was very similar to that for the GalNAc-binding mutant of the mannose-binding protein. However, the acetamido group of GalNAc appeared to be recognized more strongly by the combination of hydrogen bonds to Arg¹¹⁵ and van der Waals interaction with Gln⁷⁰. Mutational analyses, in which Gln⁷⁰ and/or Arg¹¹⁵ were replaced by alanine, confirmed that these residues contributed to GalNAc recognition in a cooperative manner.

	INTRODUCTION

TOP ABSTRACT INTRODUCTION EXPERIMENTAL PROCEDURES RESULTS AND DISCUSSION REFERENCES

 
Carbohydrate-binding proteins (lectins) have been shown to play important roles in molecular recognition processes in various tissues and body fluids. There are increasing numbers of animal lectins (1, 2), which are categorized into several families according to their structural homologies. Various proteins that display carbohydrate-binding activity in a Ca²⁺-dependent manner are classified into the C-type lectin family (3). They contain C-type carbohydrate-recognition domains (CRDs)¹ composed of 110–130 amino acid residues in common. In vertebrate C-type lectins, the C-type CRDs mostly occur as carbohydrate-binding modules linked to other domains with distinct functions. They fall into seven groups (4) and function through cooperation of the CRDs with other individual domains. In contrast, invertebrate C-type lectins are mostly single-domain proteins. One of their possible roles is thought to be inactivation or opsonization of foreign microorganisms in place of immunoglobulins in vertebrates. There are only limited numbers of C-type CRDs whose tertiary structures have been determined. Mannose-binding protein (MBP) was the first C-type lectin for which a crystal structure was determined (5). This protein contains an N-terminal collagenous domain, followed by a link domain and a C-type CRD. The latter two domains have been expressed in a recombinant form, and the structure as well as various mutant forms have been analyzed by x-ray crystallography (6–10). In addition to MBP, the crystal structures of CRDs complexed with carbohydrates have been solved for DC-SIGN, DC-SIGNR (11), P- and E-selectins (12), tunicate lectin TC14 (13), and rattlesnake venom lectin RSL (14). In addition to these lectins, several C-type lectin-like domains (CTLD) without carbohydrate-binding ability have also been found (2). These lack the essential residues for carbohydrate binding, and instead many are thought to function as receptors for noncarbohydrate ligands. Although these proteins share only slight sequence similarity (20%), their tertiary structures are basically similar to each other (15).

We previously isolated four Ca²⁺-dependent galactose/N-acetylgalactosamine-specific lectins (CEL-I, -II, -III, and -IV) from the marine invertebrate Cucumaria echinata (Holothuroidea) (16). Based on their amino acid sequences, CEL-I and CEL-IV are clearly categorized into the C-type lectin family, whereas CEL-III is a novel Ca²⁺-dependent lectin with strong hemolytic activity as well as cytotoxicity (17–20). CEL-III shows sequence similarity with -trefoil lectins, such as the B-chains of ricin and abrin (21). The similarity of the three-dimensional structure of CEL-III with these proteins has recently been revealed by x-ray crystallographic analysis (22). CEL-I, which is the smallest lectin in C. echinata, is composed of two identical 16-kDa subunits linked by a single interchain disulfide bond (23). CEL-I has very high specificity for N-acetylgalactosamine (GalNAc), and its affinity for GalNAc is 1000-fold stronger than that for galactose as judged by hemagglutination inhibition assays. To date, no other C-type lectins with such high specificity for GalNAc have been identified. It is therefore of great interest to clarify the CEL-I-specific carbohydrate-recognition mechanism not only to understand the carbohydrate-recognition mechanism of C-type lectins but also the structural basis to design molecules with novel carbohydrate-specific affinity. We previously obtained single crystals of CEL-I that were suitable for x-ray diffraction studies (24). Recently, we also succeeded in crystallizing the CEL-I-GalNAc complex. In this report, we describe x-ray crystallographic analyses of CEL-I and the CEL-I-GalNAc complex to elucidate the mechanism of its high specificity for GalNAc along with the site-directed mutagenesis experiments of a recombinant CEL-I (rCEL-I) that was expressed in Escherichia coli cells.

	EXPERIMENTAL PROCEDURES

TOP ABSTRACT INTRODUCTION EXPERIMENTAL PROCEDURES RESULTS AND DISCUSSION REFERENCES

 
Purification of CEL-I—CEL-I was purified from C. echinata according to a previously reported method (19). The proteins extracted from a homogenate of C. echinata were applied to a lactose-Cellulofine column equilibrated with 0.15 M NaCl, 10 mM Tris-HCl, pH 7.5, (TBS) containing 10 mM CaCl₂. Adsorbed lectins (CEL-I, -III, and -IV) were eluted with TBS containing 20 mM EDTA. The lectins were then separated using a GalNAc-Cellulofine column utilizing the differences in their carbohydrate-binding specificities. After elution of CEL-III with TBS containing 0.1 M lactose, CEL-I and CEL-IV were eluted with TBS containing 20 mM EDTA. CEL-I and CEL-IV were finally separated by gel filtration through Sephadex G-75 in TBS.

Crystallizations and Data Collection—The purification and crystallization of native CEL-I have already been reported (24). Briefly, the protein solution (5 mg/ml, 2–4 µl) in TBS was mixed with the same amount of reservoir solution (0.1 M Tris-HCl, pH 8.0, 60% MPD), and subjected to hanging or sitting drop vapor diffusion at 20 °C. X-ray data collection was performed on beamline BL-18B at the High Energy Acceleration Research Organization, Tsukuba, Japan, using an ADSC Quantum 4R CCD camera (25). The space group was monoclinic C2 with unit cell parameters of a = 92.4, b = 69.9, c = 76.7 Å, and = 136.5°. Reduction of a total of 72,844 reflections from CEL-I crystals yielded 19,924 independent reflections with 92.3% completeness for a resolution range of 52.7–2.0 Å and an overall Rmerge of 6.1%. CEL-I-GalNAc complex crystals were prepared under essentially the same conditions as the native crystals except for the addition of 1 mM GalNAc to the samples. Because the complex crystals were initially formed as spherulites, these were used as seeds for a microseeding method. Single crystals of the CEL-I-GalNAc complex grew to sufficient sizes for data collection within one month. Data collection was performed at 100 K using a Rigaku R-AXIS VII imaging plate area detector equipped with a Rigaku FR-E SuperBright rotating-anode generator. Image data were processed by the programs MOSFLM (26) and SCALA (27). The space group and unit cell parameters of the CEL-I/GalNAc crystals were determined to be P2₁ and a = 39.6, b = 52.2, c = 136.6, and = 91.7°, respectively. Assuming two molecules of CEL-I, each composed of a homodimer in the asymmetric unit, the value of the Matthews constant, V_m (28), is 2.20 ų Da^-1 corresponding to a solvent content of 44.2%. The final data for the CEL-I-GalNAc complex crystals comprised 59,176 independent reflections with 96.3% completeness for a range of 33.8–1.7 Å and an overall Rmerge of 6.2%. The completeness and Rmerge for the highest resolution shell (1.79–1.70 Å) were 84.9 and 16.2%, respectively.

Structure Determination and Refinement—The crystal structures of native CEL-I and the CEL-I-GalNAc complex were solved by the molecular replacement method. The search model for native CEL-I was based on the human lithostathine structure (PDB code 1LIT [PDB] ) (29), which shares 28% amino acid sequence identity with CEL-I. Molecular replacement calculations were performed using the program AMoRe (30) from the CCP4 suite (26). The model was refined using the program CNS (31) with noncrystallographic symmetry restraints. Five percent of the reflections were set aside for R_free calculations (32). Manual fitting of the model was carried out by the program Xfit (33). The final R and R_free factors for all reflections between 52.7- and 2.0-Å resolution for native CEL-I were 0.140 and 0.179, respectively. The structure of the CEL-I-GalNAc complex was also solved by the molecular replacement method using the native CEL-I coordinates as a search model. The positions and orientations of GalNAc in the complex were clearly shown by the F_o - F_c difference Fourier map with a contour level of 3. The model was finally refined by the program REFMAC (34) without noncrystallographic symmetry restraints. The final R and R_free factors between 33.8- and 1.7-Å resolution for the CEL-I-GalNAc complex were 0.158 and 0.186, respectively. The quality of the final models for native and complexed CEL-I was assessed by Ramachandran plots and analysis of the model geometry with the program PROCHECK (35). The crystallographic statistics are summarized in Table I.

Hemagglutination Assay—Serial 2-fold dilutions of a sample (50 µl) were mixed with the same volume of a 5% (v/v) suspension of rabbit erythrocytes in the wells of round bottomed microtiter plates. Incubation was performed in TBS containing 10 mM CaCl₂. The extent of agglutination was examined visually after incubation for 1 h at room temperature. The hemagglutinating activity was expressed as a titer, i.e. the reciprocal of the highest dilution producing detectable agglutination. Hemagglutination inhibition assays were performed by incubating 50-µl aliquots of the protein solutions (titer 2) containing various concentrations of carbohydrates with the same volume of a 5% (v/v) suspension of rabbit erythrocytes in TBS containing 10 mM CaCl₂.

	RESULTS AND DISCUSSION

TOP ABSTRACT INTRODUCTION EXPERIMENTAL PROCEDURES RESULTS AND DISCUSSION REFERENCES

 
The crystal structure of CEL-I was solved by the molecular replacement method using human lithostathine, which contains a CTLD (29), as a search model. Native CEL-I is composed of one disulfide-linked homodimer (protomers A and B) in each asymmetric unit (Fig. 1). As shown in other C-type CRDs, the three-dimensional structure of CEL-I consists of two parts, a lower part composed of two -helices (1 and 2) and four -strands (0, 1, 1', and 5), and an upper part composed of four -strands (2, 2', 3, and 4), the latter of which contains the calcium-binding sites (Figs. 1 and 2). Regardless of the low sequence similarity (16–30%) with other C-type CRDs and CTLDs, structural comparison with known three-dimensional structures by DALI (37) revealed that CEL-I is very similar to long form CTLDs, such as lithostathine (Z = 21.6, r.m.s.d. of 1.7 Å for 128 C atoms), the H1 subunit of the asialoglycoprotein receptor (Z = 19.6, r.m.s.d. of 1.3 Å for 118 C atoms), and tetranectin (Z = 19.5, r.m.s.d. of 2.1 Å for 132 C atoms). CEL-I also shows similarity with short form CTLDs, such as MBP-A (Z = 17.5, r.m.s.d. of 2.1 Å for 123 C atoms) and tunicate lectin TC14 (Z = 15.8, r.m.s.d. of 2.2 Å for 117 C atoms). Compared with other CRDs, CEL-I has an insertion (residues 36–41) between 1 and 1' containing a 3₁₀ helix (residues 37–39) (Fig. 3).

View larger version (37K): [in this window] [in a new window]   FIG. 1. Stereo view of the ribbon model of CEL-I (protomer A). The -helices, -strands, loops, and intramolecular disulfide bonds are shown in purple, blue, gray, and green, respectively. Bound calcium ions are shown by cyan spheres. Secondary structures were determined by the program PROMOTIF (45). Fig. 1 was drawn by the program MOLSCRIPT (46) and rendered by the program Raster3D (47).

View larger version (19K): [in this window] [in a new window]   FIG. 2. Ca2+-binding modes of CEL-I. Carbon and calcium atoms are shown in green. Oxygen and nitrogen atoms are shown in red and blue, respectively. Hydrogen and coordination bonds are indicated by dots. A, CA1 and CA3 of protomer A. The main-chain carbonyl group of the symmetry-related Gly93 in protomer B is illustrated in orange. B, CA2 of protomer A. An MPD molecule coordinating with CA2 is also shown. Hydrogen bonds between the MPD and CEL-I molecules are illustrated by dots and were determined by the program HBPLUS (48). Fig. 2 was drawn by the program PyMOL (49).

View larger version (23K): [in this window] [in a new window]   FIG. 3. Sequence alignment and secondary structure of CEL-I and other C-type lectins. Three invertebrate CRDs (CEL-I, tunicate, and starfish lectins) and a GalNAc-binding (QPDWGH) mutant of MBP (10) were aligned. The colors for the secondary structures are the same as those described in the legend to Fig. 1. Conserved residues are illustrated in red. Conserved residues among both the galactose- and mannose-binding types and the galactose-binding type only of the C-type lectins are shown in yellow and orange, respectively. Cysteine residues widely conserved among CTLDs are shown in green. The structure-based sequence alignment was performed by DALI (37), except for the GalNAc-specific binding starfish lectin. The residues that coordinate with carbohydrate-binding Ca2+ ions are boxed in blue. This figure was drawn by the program ALSCRIPT (50) and manually modified.

The two identical protomers of CEL-I are linked by an interchain disulfide bond between the Cys³⁶ residues (Fig. 4A). In addition to this disulfide bond, there are several interactions between the two subunits around the N-terminal (residues 1–4), C-terminal (residues 137–140), and insertion loop (residues 36–41) (Fig. 5). Eight hydrogen bonds are present between the two protomers (Gln²–Gln², Gln²–Gly¹¹, Glu¹⁴⁰–Thr⁵, Tyr³⁴ (protomer A)–Thr³⁸ (protomer B) and Thr³⁸ (protomer A)–Ser³⁵ (protomer B)). In particular, Gln² appears to be especially important for dimeric interactions, because the O-1 and N-2 atoms of its side chain in one subunit make hydrogen bonds to the main-chain amide group of Gln² and the main-chain carbonyl group of Gly¹¹ in the other subunit, respectively (Fig. 5A). There are also hydrophobic interactions between the two subunits through Pro⁴, Leu³⁹, Val⁴¹, Leu¹³⁷, and Phe¹³⁹ (Fig. 5B). Among the available C-type lectin crystal structures, only tunicate lectin TC14 has a homodimeric structure composed of two C-type CRDs (39). The two subunits of the TC14 homodimer associate with each other through noncovalent interactions between -strands and -helices (Fig. 4B), because two N-terminal -strands (1) make an anti-parallel -sheet, and two -helices (2) associate by hydrophobic interactions. Recently, another C-type CRD linked by intermolecular disulfide bonds has been reported in the decameric structure of rattlesnake venom lectin RSL (14). Meanwhile, the coagulation factor-binding proteins also form a disulfide-linked dimer composed of CTLDs, which have long loops to make dimeric interactions (40, 41)

View larger version (33K): [in this window] [in a new window]   FIG. 4. Homodimeric structures of CEL-I and TC14. A, CEL-I molecule. B, TC14 molecule. The colors are the same as those described in the legend to Fig. 1. In the top figures, pseudo 2-fold axes pass through the center of the two protomers perpendicular to the paper. The bottom figures were obtained by a 90-degree rotation around the horizontal line at the center of the top figures. Fig. 4 was drawn by the program MOLSCRIPT (46) and rendered by the program Raster3D (47).

View larger version (25K): [in this window] [in a new window]   FIG. 5. Dimeric interactions of CEL-I. Hydrophilic (A) and hydrophobic (B) interactions connecting the two subunits of CEL-I. Pseudo 2-fold axes pass through the center of the two protomers. Only the residues forming each hydrophilic and hydrophobic interaction are shown. The subunits in pink and blue represent protomer A and protomer B, respectively. Oxygen and nitrogen atoms of the illustrated side chains and the backbone atoms are shown in red and blue, respectively. Hydrogen bonds are indicated by yellow dots. Fig. 5 was drawn by the program PyMOL (49).

There are two CEL-I homodimer molecules (the subunit structure is designated 1A/1B and 2A/2B for each molecule) in the asymmetric unit of the CEL-I-GalNAc complex crystal. Four bound GalNAc molecules coordinate to CA2 with their 3-OH and 4-OH in place of the two hydroxyl groups of MPD found in the native CEL-I crystal (Fig. 6A). These hydroxyl groups also form hydrogen bonds with the side chains of Gln¹⁰¹, Asp¹⁰³, Glu¹⁰⁹, and Asn¹²³. This binding mode is basically consistent with those of the carbohydrate complexes of MBP-C (9) and the GalNAc-binding mutant of MBP (10) (Fig. 6B). Although the indole rings of Trp¹⁰⁵ in protomers 1B and 2B are flipped relative to those in protomers 1A and 2A, because of the hydrogen-bonding with the main-chain carbonyl oxygen of Gln¹⁰¹ in the symmetry-related molecule of protomers 1B and 2B, the Trp¹⁰⁵ residues in all protomers make van der Waals contact with the C-6 atom of GalNAc, thereby stabilizing the binding of GalNAc. Tryptophan residues have been found to stabilize the binding of carbohydrates in the Gal/GalNAc-recognizing C-type CRDs (7, 13). However, the interaction between Trp¹⁰⁵ and the C-6 atom of GalNAc appears to be relatively weak in the case of CEL-I, compared with that of Trp¹⁸⁹ in the GalNAc-binding mutant of MBP, which makes van der Waals contacts with the C-3, C-4, C-5, and C-6 carbon atoms of GalNAc (Fig. 6B). In the mutant MBP, five residues (192–196) inserted as a glycine-rich loop (7) appear to lead to closer contact between Trp¹⁸⁹ and GalNAc.

View larger version (44K): [in this window] [in a new window]   FIG. 6. Comparison of the carbohydrate-binding modes of CEL-I and the GalNAc-binding mutant of MBP. A, electron density map around the GalNAc-binding site of CEL-I (protomer 1A). A 2Fobs - Fcalc omit map is drawn at the 1.5 level. Carbon, oxygen, nitrogen, and calcium atoms are shown in green, red, blue, and purple, respectively. B, overlapped structures of CEL-I (protomer 1A) and the mutant MBP around the bound GalNAc. C, overlapped structures of CEL-I (protomer 1A) and the mutant MBP around the acetamido group. GalNAc, the calcium ion, and its binding residues in CEL-I and the MBP mutant are superimposed. Carbon, calcium atoms, and residue numbers of CEL-I and the mutant MBP are shown in green and yellow, respectively. Hydrogen and coordination bonds are indicated by dots. The GalNAc-binding site structure of protomer 2A is similar to this site. Fig. 6 was drawn by the program PyMOL (49).

CEL-I exhibits strong cytotoxicity (36), which suggests an actual biological role for this protein as a defense toxin against predators. The toxicity is inhibited in the presence of GalNAc suggesting that it is mediated by binding to specific carbohydrate chains on the cell surface. One probable mechanism is that the binding of CEL-I to cell surface carbohydrate chains triggers intracellular signaling, leading to cell death. An identification of the natural carbohydrate ligands for CEL-I that are present on the target cell surface seems to be important for understanding the cytotoxicity at the molecular level. Further investigation of the involvement of the residues around the carbohydrate-binding site of CEL-I using recombinant proteins should provide important clues regarding the recognition mechanism for natural ligands on the target cell surface.

	FOOTNOTES

 
The atomic coordinates and structure factors (codes 1WMY [PDB] and 1WMZ [PDB] ) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

^* The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^¶ Present address: Dept. of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, Komaba 3-8-1, Meguro-ku, Tokyo 153-8902, Japan.

^** To whom correspondence should be addressed. Tel.: 81-95-819-2686; Fax: 81-95-819-2684; E-mail: thata{at}net.nagasaki-u.ac.jp.

¹ The abbreviations used are: CRD, carbohydrate recognition domain; CTLD, C-type lectin-like domain; MBP, mannose-binding protein; MPD, 2-methyl-2,4-pentanediol; rCEL-I, recombinant CEL-I; DC-SIGN, dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin; DC-SIGNR, DC-SIGN-related molecule; TBS, Tris-buffered saline; r.m.s.d., root mean square deviation.

	ACKNOWLEDGMENTS

 
We thank Dr. N. Sakabe of the Foundation for Advancement of International Science and Drs. N. Watanabe, M. Suzuki, and N. Igarashi of the National Laboratory for High Energy Physics for their kind help in the data collection at the Photon Factory. This study was performed under the Cooperative Research Program of the Institute for Protein Research, Osaka University.

	REFERENCES

TOP ABSTRACT INTRODUCTION EXPERIMENTAL PROCEDURES RESULTS AND DISCUSSION REFERENCES

 

1. Gabius, H. J. (1997) Eur. J. Biochem. 243, 543-576[Medline] [Order article via Infotrieve]
2. Dodd, R. B., and Drickamer, K. (2001) Glycobiol. 11, 71R-79R[Abstract/Free Full Text]
3. Drickamer, K. (1988) J. Biol. Chem. 263, 9557-9560[Free Full Text]
4. Day, A. J. (1994) Biochem. Soc. Trans. 22, 83-88[Medline] [Order article via Infotrieve]
5. Weis, W. I., Kahn, R., Fourme, R., Drickamer, K., and Hendrickson, W. A. (1991) Science 254, 1608-1615[Abstract/Free Full Text]
6. Weis, W. I., Drickamer, K., and Hendrickson, W. A. (1992) Nature 360, 127-134[CrossRef][Medline] [Order article via Infotrieve]
7. Iobst, S. T., and Drickamer, K. (1994) J. Biol. Chem. 269, 15512-15519[Abstract/Free Full Text]
8. Kolatkar, A. R., and Weis, W. I. (1996) J. Biol. Chem. 271, 6679-6685[Abstract/Free Full Text]
9. Ng, K. K., Drickamer, K., and Weis, W. I. (1996) J. Biol. Chem. 271, 663-674[Abstract/Free Full Text]
10. Kolatkar, A. R., Leung, A. K., Isecke, R., Brossmer, R., Drickamer, K., and Weis, W. I. (1998) J. Biol. Chem. 273, 19502-19508[Abstract/Free Full Text]
11. Feinberg, H., Mitchell, D. A., Drickamer, K., and Weis, W. I. (2001) Science 294, 2163-2166[Abstract/Free Full Text]
12. Somers, W. S., Tang, J., Shaw, G. D., and Camphausen, R. T. (2000) EMBO J. 19, 467-479
13. Deleted in proof
14. Walker, J. R., Nagar, B., Young, N. M., Hirama, T., and Rini, J. M. (2004) Biochemistry 43, 3783-3792[CrossRef][Medline] [Order article via Infotrieve]
15. Swaminathan, G. J., Weaver, A. J., Loegering, D. A., Checkel, J. L., Leonidas, D. D., Gleich, G. J., and Acharya, K. R. (2001) J. Biol. Chem. 276, 26197-26203[Abstract/Free Full Text]
16. Hatakeyama, T., Kohzaki, H., Nagatomo, H., and Yamasaki, N. (1994) J. Biochem. (Tokyo) 116, 209-214[Abstract/Free Full Text]
17. Oda, T., Tsuru, M., Hatakeyama, T., Nagatomo, H., Muramatsu, T., and Yamasaki, N. (1997) J. Biochem. (Tokyo) 121, 560-567[Free Full Text]
18. Hatakeyama, T., Furukawa, M., Nagatomo, H., Yamasaki, N., and Mori, T. (1996) J. Biol. Chem. 271, 16915-16920[Abstract/Free Full Text]
19. Hatakeyama, T., Nagatomo, H., and Yamasaki, N. (1995) J. Biol. Chem. 270, 3560-3564[Abstract/Free Full Text]
20. Oda, T., Shinmura, N., Nishioka, Y., Komatsu, N., Hatakeyama, T., and Muramatsu, T. (1999) J. Biochem. (Tokyo) 125, 713-720[Abstract/Free Full Text]
21. Nakano, M., Tabata, S., Sugihara, K., Kouzuma, Y., Kimura, M., and Yamasaki, N. (1999) Biochim. Biophys. Acta 1435, 167-176[CrossRef][Medline] [Order article via Infotrieve]
22. Uchida, T., Yamasaki, T., Eto, S., Sugawara, H., Kurisu, G., Nakagawa, A., Kusunoki, M., and Hatakeyama, T. (2004) J. Biol. Chem. 279, 37133-37141[Abstract/Free Full Text]
23. Hatakeyama, T., Matsuo, N., Shiba, K., Nishinohara, S., Yamasaki, N., Sugawara, H., and Aoyagi, H. (2002) Biosci. Biotechnol. Biochem. 66, 157-163[CrossRef][Medline] [Order article via Infotrieve]
24. Hatakeyama, T., Matsuo, N., Aoyagi, H., Sugawara, H., Uchida, T., Kurisu, G., and Kusunoki, M. (2002) Acta Crystallogr. Sect. D 58, 143-144[CrossRef][Medline] [Order article via Infotrieve]
25. Watanabe, N., Nakagawa, A., Adachi, S., and Sakabe, N. (1995) Rev. Sci. Instrum. 66, 1824-1826
26. Leslie, A. G. W. (1992) Joint CCP4 + ESF-EAMCB Newsletter on Protein Crystallography 26
27. Collaborative Computational Project, Number 4 (1994) Acta Crystallogr. Sect. D 50, 760-763[CrossRef][Medline] [Order article via Infotrieve]
28. Matthews, B. W. (1968) J. Mol. Biol. 33, 491-497[Medline] [Order article via Infotrieve]
29. Bertrand, J. A., Pignol, D., Bernard, J. P., Verdier, J. M., Dagorn, J. C., and Fontecilla-Camps, J. C. (1996) EMBO J. 15, 2678-2684[Medline] [Order article via Infotrieve]
30. Navaza, J. (1994) Acta Crystallogr. Sect. A 50, 157-163[CrossRef]
31. Brunger, A. T., Adams, P. D., Clore, G. M., DeLano, W. L., Gros, P., Grosse-Kunstleve, R. W., Jiang, J. S., Kuszewski, J., Nilges, M., Pannu, N. S., Read, R. J., Rice, L. M., Simonson, T., and Warren, G. L. (1998) Acta Cryst. Sect. D 54, 905-921[CrossRef][Medline] [Order article via Infotrieve]
32. Brunger, A. (1997) Methods Enzymol. 277, 366-396[Medline] [Order article via Infotrieve]
33. McRee, D. E. (1999) J. Struct. Biol. 125, 156-165[CrossRef][Medline] [Order article via Infotrieve]
34. Murshudov, G. N., Vagin, A., and Dodson, E. (1997) Acta Crystallogr. Sect. D 53, 240-255[CrossRef][Medline] [Order article via Infotrieve]
35. Laskowski, R., MacArthur, M., Moss, D., and Thornton, J. M. (1993) J. Appl. Crystallogr. 26, 283-291[CrossRef]
36. Hatakeyama, T., Shiba, K., Matsuo, N., Fujimoto, T., Oda, T., Sugawara, H., and Aoyagi, H. (2004) J. Biochem. (Tokyo) 135, 101-107[Abstract/Free Full Text]
37. Holm, L., and Sander, C. (1994) Proteins 19, 165-173[CrossRef][Medline] [Order article via Infotrieve]
38. Drickamer, K. (1999) Curr. Opin. Struct. Biol. 9, 585-590[CrossRef][Medline] [Order article via Infotrieve]
39. Poget, S. F., Legge, G. B., Proctor, M. R., Butler, P. J., Bycroft, M., and Williams, R. L. (1999) J. Mol. Biol. 290, 867-879[CrossRef][Medline] [Order article via Infotrieve]
40. Mizuno, H., Fujimoto, Z., Koizumi, M., Kano, H., Atoda, H., and Morita, T. (1997) Nat. Struct. Biol. 4, 438-441[CrossRef][Medline] [Order article via Infotrieve]
41. Mizuno, H., Fujimoto, Z., Koizumi, M., Kano, H., Atoda, H., and Morita, T. (1999) J. Mol. Biol. 289, 103-112[CrossRef][Medline] [Order article via Infotrieve]
42. Kjell Håkansson, K., Lim, N. K., Hoppe, H. J., and Reid, K. B. (1999) Structure 7, 255-264[Medline] [Order article via Infotrieve]
43. Kakiuchi, M., Okino, N., Sueyoshi, N., Ichinose, S., Omori, A., Kawabata, S., Yamaguchi, K., and Ito, M. (2002) Glycobiology 12, 85-94[Abstract/Free Full Text]
44. Iobst, S. T., and Drickamer, K. (1996) J. Biol. Chem. 271, 6686-6693[Abstract/Free Full Text]
45. Hutchinson, E. G., and Thornton, J. M. (1996) Protein Sci. 5, 212-220[Abstract]
46. Kraulis, P. J. (1991) J. Appl. Crystallogr. 24, 946-950[CrossRef]
47. Merritt, E., and Bacon, D. (1997) Methods Enzymol. 277, 505-524[Medline] [Order article via Infotrieve]
48. McDonald, I. K., and Thornton, J. M. (1994) J. Mol. Biol. 238, 777-793[CrossRef][Medline] [Order article via Infotrieve]
49. DeLano, W. L. (2002) The PyMOL Molecular Graphics Systems, DeLano Scientific, San Carlos, CA
50. Barton, G. J. (1993) Protein Eng. 6, 37-40[Free Full Text]

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